Recombinant DNA replicable in microorganisms belonging to the genus Corynebacterium, which contains a DNA fragment coding for an aspartokinase .alpha.-subunit protein originating from a bacterium belonging to the genus Corynebacterium, in which synergistic feedback inhibition by L-lysine and L-threonine is substantially desensitized, and a DNA fragment coding for an aspartokinase .beta.-subunit protein originating from a bacterium belonging to the genus Corynebacterium, in which synergistic feedback inhibition by L-lysine and L-threonine is substantially desensitized, is introduced into a microorganism belonging to the genus Corynebacterium. Thus a transformant having enhanced production and excretion speeds of L-lysine is obtained. The transformant is cultivated in an appropriate medium, and produced L-lysine is separated.

TECHNICAL FIELD 
The present invention relates to a novel aspartokinase and a DNA fragment 
coding for the enzyme originating from a bacterium belonging to the genus 
Corynebacterium used for fermentative production of amino acid and so on, 
and relates to recombinant DNA containing the DNA fragment. The present 
invention also relates to a bacterium belonging to the genus 
Corynebacterium harboring the recombinant DNA, and relates to a method of 
producing L-lysine comprising cultivating the microorganism. 
BACKGROUND ART 
L-Lysine, which is used as a feed additive, is usually produced by 
fermentation by using an L-lysine-producing mutant strain belonging to 
coryneform bacteria. A variety of L-lysine-producing bacteria are known at 
present, which are those created by artificial mutation of coryneform 
bacteria. Such artificial mutant strains includes the followings: 
S-(2-aminoethyl)cysteine (hereinafter abbreviated as "AEC") resistant 
mutant strains; mutant strains which require amino acid such as 
L-homoserine for their growth (Japanese Patent Publication Nos. 48-28078 
and 56-6499); mutant strains which exhibit resistance to AEC and require 
amino acids such as L-leucine, L-homoserine, L-proline, L-serine, 
L-arginine, L-alanine, and L-valine (U.S. Pat. Nos. 3,708,395 and 
3,825,472); L-lysine-producing mutant strains which exhibit resistance to 
DL-.alpha.-amino-.epsilon.-caprolactam, .alpha.-amino-lauryllactam, 
aspartic acid-analog, sulfa drug, quinoid, and N-lauroylleucine; 
L-lysine-producing mutant strains which exhibit resistance to inhibitors 
of oxyaloacetate decarboxylase or respiratory system enzymes (Japanese 
Patent Laid-open Nos. 50-53588, 50-31093, 52-102498, 53-9394, 53-86089, 
55-9783, 55-9759, 56-32995 and 56-39778, and Japanese Patent Publication 
Nos. 53-43591 and 53-1833); L-lysine-producing mutant strains which 
require inositol or acetic acid (Japanese Patent Laid-open Nos. 55-9784 
and 56-8692); L-lysine-producing mutant strains which exhibit sensitivity 
to fluoropyruvic acid or temperature not less than 34.degree. C. (Japanese 
Patent Laid-open Nos. 55-9783 and 53-86090); and mutant strains belonging 
to the genus Brevibacterium or Corynebacterium which exhibit resistance to 
ethylene glycol and produce L-lysine (U.S. patent application Ser. No. 
333,455). 
Escherichia coli transformed by using a recombinant vector is disclosed in 
the prior art. In this strain, amino acid production is enhanced (see U.S. 
Pat. No. 4,278,765). 
In relation to the genera Brevibacterium and Corynebacterium, there are 
disclosed a vector plasmid which is autonomously replicable in bacterial 
cells and has a drug resistance marker gene (see U.S. patent application 
Ser. No. 386,980), and a method for introducing genes into bacterial cells 
(Japanese Patent Laid-open No. 2-207791). There is disclosed a possibility 
to breed L-threonine- or L-isoleucine-producing bacteria by using the 
techniques described above (see U.S. patent application Ser. Nos. 376,396 
and 392,145). In relation to breeding of L-lysine-producing bacteria, a 
technique is known in which a gene relevant to L-lysine biosynthesis is 
integrated into a vector plasmid to amplify it in bacterial cells (for 
example, Japanese Patent Laid-open No. 56-160997). However, no prior art 
is known in which a gene is specified for aspartokinase (hereinafter 
referred to as "AK"), a mutation point on the AK gene is elucidated so 
that feedback inhibition by L-lysine and L-threonine is substantially 
desensitized, and it is elucidated that the mutation directly relates to 
productivity of L-lysine. Although a mutant AK gene is described in a few 
cases, the mutant AK gene could not be harbored as a stable plasmid (see 
Cremer, J. et al.; Applied and Environmental Microbiology, June 1991, pp. 
1746-1752). 
An object of the present invention is to make improvement to provide 
increased production and secretion speeds of L-lysine by modifying AK as 
an important enzyme for lysine biosynthesis in microorganisms of bacteria 
belonging to the genus Corynebacterium into those having a property of 
desensitization of feedback inhibition by L-lysine and L-threonine and 
feedback inhibition by L-lysine alone, and increasing their activities. 
DISCLOSURE OF THE INVENTION 
As a result of diligent studies, the present inventors have succeeded in 
obtaining mutant AK genes from a bacterium belonging to the genus 
Corynebacterium, and completed the present invention. Namely, the present 
invention lies in an aspartokinase .alpha.-subunit protein and a DNA 
fragment coding for the protein originating from a bacterium belonging to 
the genus Corynebacterium, in which synergistic feedback inhibition by 
L-lysine and L-threonine is substantially desensitized, the protein having 
an amino acid sequence defined in SEQ ID NO: 4 in Sequence Listing, or a 
sequence in which a 279th Thr residue in the amino acid sequence defined 
in SEQ ID NO: 4 is changed to an amino acid residue other than Ala and 
other than acidic amino acids. Further, the present invention lies in an 
aspartokinase .beta.-subunit protein and a DNA fragment coding for the 
protein originating from a bacterium belonging to the genus 
Corynebacterium, in which synergistic feedback inhibition by L-lysine and 
L-threonine is substantially desensitized, the protein having an amino 
acid sequence defined in SEQ ID NO: 6 in Sequence Listing, or a sequence 
in which a 30th Thr residue in the amino acid sequence defined in SEQ ID 
NO: 6 is changed to an amino acid residue other than Ala and other than 
acidic amino acids. 
In another aspect, the present invention lies in recombinant DNA containing 
the aforementioned DNA fragment and being replicable in microorganisms 
belonging to the genus Corynebacterium, and a transformant obtained by 
introducing the recombinant DNA into a microorganism belonging to the 
genus Corynebacterium, wherein the specific activity of aspartokinase is 
increased 2-20 times as compared with a parent strain, and synergistic 
feedback inhibition by L-lysine and L-threonine or feedback inhibition by 
L-lysine alone exerted on the activity of aspartokinase is substantially 
desensitized. 
In another aspect, the present invention lies in a method of producing 
L-lysine comprising the steps of cultivating the aforementioned 
transformant in an appropriate medium, and separating produced L-lysine. 
The microorganisms belonging to the genus Corynebacterium referred to in 
the present invention are a group of microorganisms as defined in Bergey's 
Manual of Determinative Bacteriology, 8th ed., p. 599 (1974), which are 
aerobic gram-positive rods having no acid resistance and no spore-forming 
ability. The microorganisms belonging to the genus Corynebacterium 
referred to in the present invention include bacteria belonging to the 
genus Brevibacterium having been hitherto classified into the genus 
Brevibacterium but united as bacteria belonging to the genus 
Corynebacterium at present, and include bacteria belonging to the genus 
Brevibacterium closely relative to bacteria belonging to the genus 
Corynebacterium. Among the microorganisms belonging to the genus 
Corynebacterium (Brevibacterium) as described above, especially glutamic 
acid-producing bacteria belonging to the genus Corynebacterium 
(Brevibacterium) as mentioned below are most preferable in the present 
invention. Further, some bacteria belonging to the genus Microbacterium 
are known to accumulate glutamic acid, which can be also used in the 
present invention. 
Examples of wild type strains of glutamic acid-producing bacteria belonging 
to the genus Corynebacterium (Brevibacterium) include the followings. 
______________________________________ 
Corynebacterium acetoacidophilum 
ATCC 13870 
Corynebacterium acetoglutamicum 
ATCC 15806 
Corynebacterium callunae 
ATCC 15991 
Corynebacterium glutamicum 
ATCC 13032 
ATCC 13060 
(Brevibacterium divaricatum) 
ATCC 14020 
(Brevibacterium lactofermentum) 
ATCC 13869 
Corynebacterium lilium ATCC 15990 
Corynebacterium melassecola 
ATCC 17965 
Brevibacterium saccharolyticum 
ATCC 14066 
Brevibacterium immariophilum 
ATCC 14068 
Brevibacterium roseum ATCC 13825 
Brevibacterium flavum ATCC 13826 
Brevibacterium thiogenitalis 
ATCC 19240 
Microbacterium ammoniaphilum 
ATCC 15354 
______________________________________ 
The glutamic acid-producing bacteria belonging to the genus Corynebacterium 
(Brevibacterium) of the present invention also include mutant strains 
having glutamic acid productivity or having lost glutamic acid 
productivity, in addition to the wild type strains having glutamic acid 
productivity as described above. 
When a wild type strain is used as a donor for the DNA fragment containing 
the AK gene, a DNA fragment containing a wild type AK gene can be 
obtained. The DNA fragment containing the gene for AK in which synergistic 
feedback inhibition by L-lysine and L-threonine is substantially 
desensitized can be obtained by using a mutant strain in which synergistic 
feedback inhibition on the AK activity by L-lysine and L-threonine is 
substantially desensitized. The mutant strain can be obtained, for 
example, from a group of cells subjected to an ordinary mutation 
treatment, ultraviolet light irradiation, or a treatment with a mutating 
agent such as N-methyl-N'-nitro-N-nitrosoguanidine (NTG). The AK activity 
can be measured by using a method described in Miyajima, R. et al., The 
Journal of Biochemistry (1968), 63(2), 139-148. 
As for the donor for the DNA fragment containing the AK gene, 
Corynebacterium glutamicum (Brevibacterium lactofermentum) wild type 
strain ATCC 13869, and an L-lysine-producing bacterium AJ3463 (FERM 
P-1987) derived by a mutation treatment from the ATCC 13869 strain are 
most preferable donors. The wild type AK gene, and the gene coding for 
aspartokinase in which synergistic feedback inhibition by L-lysine and 
L-threonine is substantially desensitized (hereinafter referred to as 
"mutant AK gene") are separated from chromosomal DNA of these bacteria, 
ligated with a vector autonomously replicable in bacteria belonging to the 
genus Corynebacterium (Brevibacterium), and introduced into cells of 
bacteria belonging to the genus Corynebacterium (Brevibacterium). 
A method for isolating the AK gene is as follows. At first, a chromosomal 
gene is extracted from a strain having the AK gene of a bacterium 
belonging to the genus Corynebacterium (for example, a method of H. Saito 
and K. Miura, Biochem. Biophys. Acta, 72, 619 (1963) can be used), and it 
is digested with a suitable restriction enzyme. Subsequently, it is 
ligated with a vector replicable in cells of bacteria belonging to the 
genus Corynebacterium. An obtained recombinant vector is used to transform 
an AK-deficient mutant strain of a microorganism belonging to the genus 
Corynebacterium. A bacterial stain consequently harboring the AK-producing 
activity is isolated, from which the AK gene can be separated. A method 
for deriving the AK-deficient mutant strain is similar for practice to a 
method for deriving the mutant strain which provides substantially 
desensitized synergistic feedback inhibition on the AK activity by 
L-lysine and L-threonine described above. 
Upon digestion of the chromosomal gene, the degree of digestion can be 
controlled by controlling the digestion reaction time and so on. Thus a 
wide variety of restriction enzymes can be used. 
Any vector replicable in cells of bacteria belonging to the genus 
Corynebacterium can be used as the vector for the present invention. 
Specifically, it is exemplified by the followings. 
(1) pAM330: see Japanese Patent Laid-open No. 58-67699 
(2) pHM1519: see Japanese Patent Laid-open No. 58-77895 
(3) pAJ655: see Japanese Patent Laid-open No. 58-192900 
(4) pAJ611: see the same 
(5) pAJ1844: see the same 
(6) pCGi: see Japanese Patent Laid-open No. 57-134500 
(7) pCG2: see Japanese Patent Laid-open No. 58-35197 
(8) pCG4: see Japanese Patent Laid-open No. 57-183799 
(9) pCG11: see the same 
The vector may be cleaved by digestion with a restriction enzyme which 
digests the DNA at one place, or by partial digestion with a restriction 
enzyme which digests it at a plurality of places. 
The vector is digested with a restriction enzyme used to digest the 
chromosomal gene, or ligated with oligonucleotides having nucleotide 
sequences complementary to both ends of the digested fragment of 
chromosomal DNA and the digested vector respectively. Subsequently, it is 
subjected to a ligation reaction of the vector and the chromosomal DNA 
fragment. 
Recombinant DNA constructed by ligating the chromosomal DNA with the vector 
thus obtained can be introduced into a recipient of bacteria belonging to 
the genus Corynebacterium by using a method in which permeability of DNA 
is increased by treating recipient cells with calcium chloride as reported 
for Escherichia coli K-12 (Mandel, M. and Higa, A., J. Mol. Biol., 53, 159 
(1970)), or by using a method in which introduction is performed in a 
proliferating stage (so-called competent cells) so that cells can 
incorporate DNA as reported for Bacillus subtills (Duncan, C. H., Wilson, 
G. A. and Young, F. E., Gene, 1, 153 (1977)). Alternatively, recombinant 
DNA can be introduced into a DNA recipient after converting DNA recipient 
cells into protoplasts or spheroplasts which easily incorporate 
recombinant DNA as known for Bacillus subtills, actinomycetes, and yeasts 
(Chang, S. and Choen, S. N., Molec. Gen. Genet., 168, 111 (1979); Bibb, M. 
J., Ward, J. M. and Hopwood, O. A., Nature, 274, 398 (1978); Hinnen, A., 
Hicks, J. B. and Fink, G. R., Proc. Natl. Acad. Sci. U.S.A., 75, 1929 
(1978)). 
In the protoplast method, a sufficiently high frequency can be obtained 
even by using the method used for Bacillus subtills described above. It is 
of course possible to utilize a method in which DNA is incorporated into 
protoplasts of a bacterium belonging to the genus Corynebacterium in the 
presence of polyethylene glycol or polyvinyl alcohol and divalent metal 
ion as described in Japanese Patent Laid-open No. 57-183799. An equivalent 
result is obtained even by using a method in which incorporation of DNA is 
facilitated by addition of carboxymethyl cellulose, dextran, Ficoll, 
Bruronik F68 (Serva) instead of polyethylene glycol or polyvinyl alcohol. 
Alternatively, the AK gene can be obtained by amplifying the AK gene by 
using PCR (polymerase chain reaction; see White, T. J. et al., Trends 
Genet., 5, 185 (1989)) from chromosomal DNA obtained as described above. 
DNA primers to be used for the amplification are those complementary to 
both 3' ends of DNA double strands containing an entire or partial region 
of the AK gene. When only a partial region of the AK gene is amplified, it 
is necessary to perform screening from a gene library by using DNA 
fragments of the region as primers to amplify a DNA fragment containing an 
entire region. When an entire region is amplified, a DNA fragment 
containing the AK gene can be recovered by excising an objective band 
after subjecting the DNA fragment to agarose gel electrophoresis. 
For DNA primers, single strand DNA's of 23 mer and 21 mer having sequences 
of 5'-TCGCGAAGTAGCACCTGTCACTT-3' (SEQ ID NO:15) and 
5'-ACGGAATTCAATCTTACGGCC-3' (SEQ ID NO:16) are most suitable to amplify a 
region of about 1,643 bp coding for the AK gene based on, for example, a 
sequence known for Corynebacterium glutamicum (see Molecular Microbiology 
(1991), 5(5), 1197-1204; Mol. Gen. Genet. (1990), 224, 317-324). The DNA 
can be synthesized in accordance with an ordinary method using a 
phosphoramidite method (see Tetrahedron Letters (1981), 22, 1859) by using 
a DNA synthesizer Model 380B produced by Applied Biosystems. The PCR can 
be performed by using DNA Thermal Cycler Model PJ2000 produced by Takara 
Shuzo Co., Ltd., using Taq DNA polymerase in accordance with a method 
designated by the supplier. 
The amplified AK gene is ligated with the vector proliferative in cells of 
bacteria belonging to the genus Corynebacterium as described above, and 
introduced into cells of bacteria belonging to the genus Corynebacterium 
by using the method as described above. 
Hosts in which the obtained AK gene is introduced and amplified to produce 
lysine include the wild type strains of glutamic acid-producing bacteria 
belonging to the genus Corynebacterium described above. All bacteria other 
than the above can be utilized as a host provided that a replication 
origin for the recombinant DNA constructed herein and the mutant AK gene 
make their function, the recombinant DNA can be replicated, and the mutant 
AK activity can be enhanced. The most preferable host is AJ12036 strain 
(FERM P-7559) which is a wild type strain of Corynebacterium glutamicum 
(Brevibacterium lactofermentum). 
The transformant harboring the recombinant DNA containing the gene coding 
for aspartokinase in which synergistic feedback inhibition by L-lysine and 
L-threonine is substantially desensitized obtained by the method described 
above is cultivated, and objective L-lysine is produced and accumulated in 
a culture liquid to collect it. 
The medium to be used for L-lysine production is an ordinary medium 
containing a carbon source, a nitrogen source, inorganic ions and 
optionally other organic components. 
As the carbon source, it is possible to use sugars such as glucose, 
lactose, galactose, fructose, and starch hydrolysate; alcohols such as 
glycerol and sorbitol; or organic acids such as fumaric acid, citric acid 
and succinic acid. 
As the nitrogen source, it is possible to use inorganic ammonium salts such 
as ammonium sulfate, ammonium chloride and ammonium phosphate; organic 
nitrogen such as soybean hydrolysate; ammonia gas; and aqueous ammonia. 
It is desirable to allow required substances such as vitamin B.sub.1 and 
L-homoserine or yeast extract to be contained in appropriate amounts as 
organic trace nutrient sources. Other than the above, potassium phosphate, 
magnesium sulfate, iron ion, manganese ion and so on are added in small 
amounts, if necessary. 
Cultivation is preferably carried out under an aerobic condition for 16-72 
hours. The cultivation temperature is controlled at 30.degree. C. to 
45.degree. C., and pH is controlled at 5-7 during cultivation. Inorganic 
or organic, acidic or alkaline substances as well as ammonia gas or the 
like can be used for pH adjustment. Collection of L-lysine from a 
cultivated liquor can be carried out by combining an ordinary ion exchange 
resin method, a precipitation method, and other known methods.

BEST MODE FOR CARRYING OUT THE INVENTION 
The present invention will be concretely explained below with reference to 
Examples. 
Example 1 
Preparation of Wild Type and Mutant AK Genes, and Preparation of Plasmids 
for Corynebacterium 
Chromosomal DNA was prepared in accordance with an ordinary method from 
Corynebacterium glutamicum (Brevibacterium lactofermentum) wild type 
strain ATCC 13869, and L-lysine-producing mutant strain AJ3463 (FERM 
P-1987) obtained therefrom by a mutation treatment. AK genes were 
amplified from the chromosomal DNA by PCR (polymerase chain reaction; see 
White, T. J. et al., Trends Genet., 5, 185 (1989)). For DNA primers used 
for the amplification, single strand DNA's of 23 mer and 21 mer having 
sequences of 5'-TCGCGAAGTAGCACCTGTCACTT-3' (SEQ ID NO: 15) and 
5'-ACGGAATTCAATCTTACGGCC-3' (SEQ ID NO: 16) were synthesized to amplify a 
region of about 1,643 bp coding for the AK genes based on a sequence known 
for Corynebacterium glutamicum (see Molecular Microbiology (1991), 5(5), 
1197-1204; Mol. Gen. Genet. (1990), 224, 317-324). The DNA was synthesized 
in accordance with an ordinary method using a phosphoramidite method (see 
Tetrahedron Letters (1981), 22, 1859) by using a DNA synthesizer Model 
380B produced by Applied Biosystems. In the PCR, the gene was amplified by 
using DNA Thermal Cycler Model PJ2000 produced by Takara Shuzo Co., Ltd., 
using Taq DNA polymerase in accordance with a method designated by the 
supplier. The amplified gene fragment of 1,643 bp was confirmed by agarose 
gel electrophoresis. Subsequently, the fragment excised from the gel was 
purified in accordance with an ordinary method, and digested with 
restriction enzymes NruI (produced by Takara Shuzo) and EcoRI (produced by 
Takara Shuzo). pHSG399 (see Takeshita, S. et al., Gene (1987), 61, 63-74) 
was used as a vector for cloning the gene fragment. pHSG399 was digested 
with restriction enzymes SmaI (produced by Takara Shuzo) and EcoRI, and 
ligated with the amplified AK gene fragment. Ligation of DNA was performed 
by using DNA ligation kit (produced by Takara Shuzo) in accordance with a 
designated method. Thus plasmids were prepared in which the AK gene 
fragments amplified from chromosome of Brevibacterium were ligated with 
pHSG399. A plasmid having the AK gene originating from ATCC 13869 as a 
wild type strain was designated as p399AKY, and a plasmid having the AK 
gene originating from AJ3463 as an L-lysine-producing bacterium was 
designated as p399AK9. 
A DNA fragment having an ability to allow plasmids to be autonomously 
proliferative in bacteria belonging to the genus Corynebacterium 
(hereinafter referred to as "Coryne.-ori") was introduced into p399AKY and 
p399AK9 respectively to prepare plasmids carrying the AK genes 
autonomously replicable in bacteria belonging to the genus 
Corynebacterium. In order to obtain Coryne.-ori, a plasmid vector pHK4 
autonomously proliferative in bacterial cells of both Escherichia coli and 
bacteria belonging to the genus Corynebacterium was prepared. Some plasmid 
vectors autonomously proliferative in bacterial cells of both Escherichia 
coli and bacteria belonging to the genus Corynebacterium had been 
reported. Herein a novel shuttle vector pHK4 was constructed from pAJ1844 
(see Japanese Patent Laid-open No. 58-216199) and pHSG298 (see S. 
Takeshita et al., Gene, 61, 63-74 (1987)). pAJ1844 was partially digested 
with a restriction enzyme Sau3AI, and ligated with pHSG298 having been 
completely digested with a restriction enzyme BamHI. Corynebacterium 
glutamicum (Brevibacterium lactofermentum) AJ12036 (FERM P-7559) was 
transformed with DNA after the ligation. An electric pulse method (see 
Japanese Patent Laid-open No. 2-207791) was used for transformation. 
Transformants were selected on M-CM2G plates containing 25 .mu.g/ml of 
kanamycin (containing glucose 5 g, polypeptone 10 g, yeast extract 10 g, 
NaCi 5 g, DL-methionine 0.2 g, and agar 15 g in 1 l of pure water, pH 
7.2). Plasmids were prepared from the transformants. One having the 
smallest size was selected, and designated as pHK4. This plasmid is 
capable of autonomous proliferation in Escherichia coli and bacteria 
belonging to the genus Corynebacterium, and gives kanamycin resistance to 
the host. 
pHK4 was digested with a restriction enzyme KpnI (produced by Takara 
Shuzo), and digested ends were blunt-ended. Blunt ends were formed by 
using DNA Blunting kit (produced by Takara Shuzo) in accordance with a 
designated method. After the blunt end formation, a phosphorylated BamHI 
linker (produced by Takara Shuzo) was ligated to make modification so that 
a DNA fragment of the Coryne.-ori portion could be excised from pHK4 by 
digestion only with BamHI. This plasmid was digested with BamHI. A 
generated Coryne.-ori DNA fragment was ligated with p399AKY and p399AK9 
having been also digested with BamHI to prepare plasmids being 
autonomously proliferative in bacteria belonging to the genus 
Corynebacterium and containing the AK genes. The plasmid containing the 
wild type AK gene originating from p399AKY was designated as p399AKYB, and 
the plasmid containing the mutant AK gene originating from p399AK9 was 
designated as p399AK9B. Steps of constructing p399AK9B and p399AKYB are 
shown in FIG. 1. A strain AJ12691 obtained by introducing the mutant AK 
plasmid p399AK9B into AJ12086 strain (FERM P-7559) as a wild type strain 
of Corynebacterium glutamicum (Brevibacterium divaricatum) has been 
awarded a deposition number (FERM P-12918), and deposited in National 
Institute of Bioscience and Human Technology of Agency of Industrial 
Science and Technology. 
Example 2 
Determination of Nucleotide Sequences of Wild Type AK and Mutant AK Genes 
of Corynebacterium glutamicum 
The plasmid p399AKY containing the wild type AK gene, and the plasmid 
p299AK9 containing the mutant AK gene were prepared to determine 
nucleotide sequences of the wild type and mutant AK genes. Nucleotide 
sequences were determined in accordance with a method of Sanger (F. Sanger 
et al., Proc. Natl. Acad. Sci., 74, 5463 (1977), etc.). The nucleotide 
sequence of the wild type AK gene encoded by p399AKY is shown in SEQ ID 
NO: 1 in Sequence Listing. The nucleotide sequence of the mutant AK gene 
encoded by p399AK9 is shown in SEQ ID NO: 2 in Sequence Listing. The 
mutant AK gene has mutation of only one base such that 1051th G is changed 
to A, as compared with the wild type AK. It is known for the AK gene that 
two subunits of .alpha. and .beta. are encoded on an identical DNA strand 
in an identical reading frame (see Kalinowski, J. et al., Molecular 
Microbiology (1991) 5(5), 1197-1204). Judging from homology, it is also 
assumed for the genes of the present invention that two subunits of 
.alpha. and .beta. are encoded on an identical DNA strand in an identical 
reading frame. 
An amino acid sequence of the .alpha.-subunit of the wild type AK protein 
deduced from the nucleotide sequence of DNA is shown in SEQ ID NO: 3 in 
Sequence Listing. An amino acid sequence of the .beta.-subunit is shown in 
SEQ ID NO: 5 in Sequence Listing. As those simultaneously showing the DNA 
sequence and the amino acid sequence, the .alpha.-subunit is shown in SEQ 
ID NO: 7 in Sequence Listing, and the .beta.-subunit is shown in SEQ ID 
NO: 9 in Sequence Listing. Nucleotide sequences corresponding to open 
reading frame portions of each of the .alpha.- and .beta.-subunits are 
shown in SEQ ID NOS: 11 and 13 in Sequence Listing, respectively. 
Similarly, an amino acid sequence of the .alpha.-subunit of the mutant AK 
protein deduced from the nucleotide sequence of DNA is shown in SEQ ID NO: 
4 in Sequence Listing. An amino acid sequence of the .beta.-subunit is 
shown in SEQ ID NO: 6 in Sequence Listing. As those simultaneously showing 
the DNA sequence and the amino acid sequence, the .alpha.-subunit is shown 
in SEQ ID NO: 8 in Sequence Listing, and the .beta.-subunit is shown in 
SEQ ID NO: 10 in Sequence Listing. Nucleotide sequences corresponding to 
open reading frame portions of each of the .alpha.- and .beta.-subunits 
are shown in SEQ ID NOS: 12 and 14 in Sequence Listing, respectively. 
In each of the subunits, GTG is used as a start codon, and a corresponding 
amino acid is depicted as methionine. However, it is intended to represent 
methionine, valine, or formylmethionine. The mutation point of the mutant 
AK Gene implies that the mutant AK protein undergoes amino acid 
substitution such that 279th alanine is changed to threonine in the 
.alpha.-subunit, and 30th alanine is changed to threonine in the 
.beta.-subunit. 
Example 3 
Effect on L-Lysine Productivity by Introduction of Mutant AK and Wild Type 
AK Plasmids into Corynebacterium glutamicum Wild Type Strain 
Strains were prepared by introducing the wild type AK plasmid p399AKYB and 
the mutant AK plasmid p399AK9B respectively into AJ12036 strain (FERM 
P-7559) as a wild type strain of Corynebacterium glutamicum 
(Brevibacterium lactofermentum). The genes were introduced into 
Corynebacterium by means of an electric pulse method. The aspartokinase 
activity was measured for Corynebacterium glutamicum (Brevibacterium 
lactofermentum) AJ12036 strain as a host, AJ12690 strain harboring the 
wild type AK plasmid, and AJ12691 (FERM P12918) strain harboring the 
mutant AK plasmid. The activity was measured in accordance with an 
ordinary method (see Miyajima, R. et al., The Journal of Biochemistry 
(1968) 63(2), 139-148). As shown in Table 1, it was confirmed that the 
introduction of the AK plasmid increased the specific activity of AK about 
10-15 times, and that the synergistic inhibition by L-lysine and 
L-threonine was desensitized only in the strain introduced with the mutant 
AK plasmid. Table 1 shows specific activities of aspartokinases of 
cell-disrupted solutions prepared from wild type AJ12036 strain of 
Corynebacterium glutamicum (Brevibacterium lactofermentum), AJ12690 strain 
additionally harboring the wild type AK plasmid, and AJ12691 strain 
additionally harboring the mutant AK plasmid, and degrees of synergistic 
inhibition thereof by L-lysine and L-threonine. L-Lysine and L-threonine 
as inhibitors were added to give a final concentration of 1 mM, 
respectively. 
TABLE 1 
______________________________________ 
AK specific activity (mU/mg protein) 
Bacterial strain 
No addition 
+1 mM L-Lys, +1 mM L-Thr 
______________________________________ 
AJ12036 19.0 2.6 
AJ12690 235.3 34.5 
AJ12691 210.5 145.3 
______________________________________ 
The lysine productivity was evaluated by cultivation for the wild type 
strain AJ12036, the wild type AK plasmid-harboring strain AJ12690, and the 
mutant AK plasmid-harboring strain AJ12691 (FERM P-12918). The evaluation 
by cultivation was performed by inoculating the strains to a production 
medium (containing glucose 100 g, (NH.sub.4).sub.2 SO.sub.4 55 g, KH.sub.2 
PO.sub.4 1 g, MgSO.sub.4.7H.sub.2 O 1 g, soybean protein acid hydrolysate 
"Mamenou" (product of Ajinomoto Co., Ltd., trade name) 50 ml, 
FeSO.sub.4.7H.sub.2 O 10 mg, MnSO.sub.4.4H.sub.2 O 10 mg, nicotinic acid 
amide 5 mg, and CaCO.sub.3 50 g in 1 l of pure water, pH 8.0), and 
cultivating them with reciprocatory shaking at 31.5.degree. C. for 72 
hours. Amounts of produced lysine in culture liquids after the cultivation 
were as shown in Table 2. It is understood that the L-lysine productivity 
is remarkably improved in the strain introduced with the mutant AK 
plasmid. The plasmid-holding ratio upon completion of the cultivation was 
measured by using an index of chloramphenicol resistance as a drug 
resistance marker of the plasmid. It was about 100%, exhibiting extremely 
high stability of the plasmid (Table 2). Table 2 shows results of 
measurement of the L-lysine productivity by fermentation and the 
plasmid-holding ratio upon completion of the cultivation for wild type 
AJ12036 strain of Corynebacterium glutamicum (Brevibacterium 
lactofermentum), AJ12690 strain additionally harboring the wild type AK 
plasmid, and AJ12691 strain additionally harboring the mutant AK plasmid. 
TABLE 2 
______________________________________ 
Amount of accumulated 
Plasmid-holding 
Bacterial strain 
Lys (g/l) ratio (%) 
______________________________________ 
AJ12036 0 -- 
AJ12690 2 100 
AJ12691 25 98 
______________________________________ 
--: no available data 
Example 4 
Analysis of Enzymes of Wild Type AK and Mutant AK of Corynebacterium 
glutamicum 
For measurement and evaluation of the enzyme activity of AK, an 
AK-completely deficient strain of Escherichia coli Gif106M1 was used as a 
host (Boy, E. and Patte, J. C., J. Bacteriol. 112, 84-92 (1972), Theze, J. 
et al., J. Bacteriol. 117, 133-143 (1974)), otherwise AK of a host and AK 
from a plasmid might exist in a mixed manner, resulting in impossibility 
to perform correct measurement, because no AK-deficient strain was present 
in bacteria belonging to the genus Corynebacterium. It is known that most 
of genes of bacteria belonging to the genus Corynebacterium are expressed 
in Escherichia coli. In addition, the AK gene was ligated at a position 
downstream from lac promoter on pHSG399. Thus it was postulated that the 
gene could be expressed in Escherichia coli. 
At first, Gif106M1 was transformed with p399AKY and p399AK9 prepared in 
Example 1 to confirm growth complementation in a minimum medium M9 shown 
below. Thus it was confirmed that AK from the bacteria belonging to the 
genus Corynebacterium was expressed and operated in cells of Escherichia 
coli. 
Minimum medium M9 
A: 20.times.M9 
Na.sub.2 HPO.sub.4.12H.sub.2 O 303 (g/L) 
KH.sub.2 PO.sub.4 60 
NaCl 10 
NH.sub.4 Cl 20 
B: 1M MgSO.sub.4 
C: 50% Glucose 
D: 1 g/L Thiamine 
A, B, C, and D were sterilized separately, and mixed in a ratio of 
A:B:C:D=5:0.1:1:0.1:95. 
Subsequently, cell-free extracts were prepared from bacterial cells, and 
the enzyme activity of AK was measured. 
Upon measurement of the enzyme activity of AK, various concentrations of 
lysine and threonine were added to enzyme reaction solutions to 
investigate the degree of inhibition (FIG. 2). As a result, it was found 
that the mutant AK exhibited little improvement in inhibition by lysine 
alone as compared with the wild type, however, inhibition by threonine was 
desensitized by 100% even with activation a little, and that the 
desensitization of inhibition by threonine resulted in mitigation of 
concerted inhibition by lysine+threonine. (Ki value: 0.4 mM.fwdarw.5.0 
mM). 
Example 5 
Preparation of Inhibition-Desensitized Type AK Gene of Corynebacterium 
glutamicum 
It was revealed from Example 4 that the mutant AK was insufficient in 
desensitization of inhibition by lysine alone. Thus it was intended to 
improve this property by introducing mutation. 
Site-directed mutagenesis was used as a method for preparing 
inhibition-desensitized type AK genes. It was intended to substitute the 
mutation point specified in Example 2 (.sup.279 Ala.fwdarw.Thr) with other 
amino acid residues. The site-directed mutagenesis method for causing 
objective mutation at an objective site includes, for example, a method 
using PCR (Higuchi, R., 61, in PCR Technology (Erlich, H. A. Eds., 
Stockton press (1989)), and a method using phage (Kramer, W. and Frits, H. 
J., Meth. in Enzymol., 154 350 (1987); Kunkel, T. A. et al., Meth. in 
Enzymol., 154 367 (1987)). 
Types of amino acid residues to be introduced by mutation were as follows. 
Twenty kinds of amino acids were classified according to several 
properties such as polarity and molecular structure, and 8 kinds of 
representatives (Arg, Asp, Cys, Phe, Pro, Ser, Tyr, Val) were selected. 
Amino acid mutation and nucleotide substitution at the mutation point of 
each of them are shown in Table 3. 
TABLE 3 
______________________________________ 
Name of plasmid 
(name after 
Name of Mutation point and 
introduction of 
mutation amino acid change 
Coryne.-ori) 
______________________________________ 
Wild type p399AKY (P399AKYB) 
Thr .sup.279 Ala GCT .fwdarw. Thr A*CT 
p399AK9 (p399AK9B) 
Arg .sup.279 Ala GCT .fwdarw. Arg C*G*T 
p399AKAR (p399AKARB) 
Asp .sup.279 Ala GCT .fwdarw. Asp GA*T 
p399AKAD 
Cys .sup.279 Ala GCT .fwdarw. Cys T*G*T 
p399AKAC (p399AKACB) 
Phe .sup.279 Ala GCT .fwdarw. Phe T*T*T 
p399AKAF (p399AKAFB) 
Pro .sup.279 Ala GCT .fwdarw. Pro C*CT 
p399AKAP 
(p399AKAPB) 
Ser .sup.279 Ala GCT .fwdarw. Ser T*CT 
p399AKAS 
(p399AKASB) 
Tyr .sup.279 Ala GCT .fwdarw. Tyr T*A*T 
p399AKAY (p399AKAYB) 
Val .sup.279 Ala GCT .fwdarw. Val GT*T 
p399AKAV 
(p399AKAVB) 
______________________________________ 
A method for introducing mutation was as follows. Eight species of 
synthetic DNA of 23 mer were designed, in which the codon of the 279th Ala 
residue for introducing mutation was substituted with the codons of the 
objective amino acid residues. Synthetic DNA for introducing Arg was 
5'-GCCAGGCGAG CGT GCCAAGGTTT-3' (SEQ ID NO: 17), synthetic DNA for 
introducing Asp was 5'-GCCAGGCGAG GAT GCCAAGGTTT-3' (SEQ ID NO: 18), 
synthetic DNA for introducing Cys was 5'-GCCAGGCGAG TGT GCCAAGGTTT-3' (SEQ 
ID NO: 19), synthetic DNA for introducing Phe was 5'-GCCAGGCGAG TTT 
GCCAAGGTTT-3' (SEQ ID NO: 20), synthetic DNA for introducing Pro was 
5'-GCCAGGCGAG CCT GCCAAGGTTT-3' (SEQ ID NO: 21), synthetic DNA for 
introducing Ser was 5'-GCCAGGCGAG TCT GCCAAGGTTT-3' (SEQ ID NO: 22), 
synthetic DNA for introducing Tyr was 5'-GCCAGGCGAG TAT GCCAAGGTTT-3' (SEQ 
ID NO: 23), and synthetic DNA for introducing Val was 5'-GCCAGGCGAG GTT 
GCCAAGGTTT-3' (SEQ ID NO: 24). Sixteen species of 23 mer single strand 
DNA's were synthesized together with complementary sequences thereof. For 
example, when the Arg residue was introduced, the single strand DNA having 
the sequence of 5'-GCCAGGCGAG CGT GCCAAGGTTT-3'(SEQ ID NO:17), the single 
strand DNA complementary thereto, the single strand DNA having the 
sequence of SEQ ID NO: 15, and the single strand DNA having the sequence 
of SEQ ID NO: 16 were used as primers to conduct the PCR method by using 
pB99AKY as a template. In order to exclude introduction of non-specific 
mutation, about 280 base pairs containing the mutation point were excised 
from prepared DNA by using restriction enzymes (NaeI-AvaII) to make 
substitution with a corresponding portion of p399AKY. The nucleotide 
sequence was confirmed for the substituted region. 
Example 6 
Analysis of 8 Kinds of Enzymes of Mutant AK Genes 
Gif106M1 was transformed with 8 kinds of the plasmids containing each of 
the mutant AK genes obtained in Example 5 in accordance with a method 
similar to that in Example 4. Cell-free extracts were prepared, and 
enzymes were analyzed. The degree of desensitization of inhibition, and 
the specific activity upon addition of lysine 5 mM, threonine 5 mM, or 
lysine 5 mM+threonine 5 mM are shown in Table 4. The degree of 
desensitization of inhibition upon addition of each concentration of 
lysine and/or threonine is graphically shown in FIGS. 3, 4 and 5. 
TABLE 4 
______________________________________ 
Specific 
activity 
(mU/mg 
5 mM Lys 5 mM Thr 5 mM Lys 
protein) 
(%) (%) + Thr (%) 
______________________________________ 
Wild type 15.3 42.3 62.3 9.2 
Thr 12.9 47.0 103.5 50.4 
Pro 2.8 76.9 126.9 103.8 
Cys 15.4 56.3 108.1 17.0 
Ser 8.2 52.6 131.6 18.4 
Val 21.8 51.1 98.3 52.3 
Arg 7.6 40.6 107.2 47.8 
Tyr 14.4 14.4 103.6 19.4 
Phe 18.7 12.1 103.0 18.2 
Asp 1.5 -- -- -- 
______________________________________ 
AK was inactivated in the case of change to acidic amino acid such as Asp. 
However, the inhibition by threonine was desensitized in the case of 
change to any other amino acid. The mutation could be generally classified 
into 4 groups for other properties. Mutation similar to the mutant (Thr) 
in Example 2 includes the Val residue-introduced mutant strain, and the 
Arg residue-introduced mutant strain. As for the Cys residue-introduced 
mutant strain and the Ser residue-introduced mutant strain, the inhibition 
by lysine alone was equivalent to that in the wild type, however, the 
inhibition was consequently enhanced in the case of the concerted 
inhibition. The concerted inhibition has been enhanced probably because 
the behavior to threonine has a characteristic property to give a 
crest-shaped graph such that activation occurs at low concentrations but 
inhibition occurs at high concentrations. The inhibition by lysine alone 
Was enhanced in the Phe residue-introduced mutant strain and the Tyr 
residue-introduced mutant strain concerning aromatic amino acids, as 
compared with the wild type. The Pro residue-introduced mutant strain had 
a low specific activity (about 1/5 of that of the wild type) probably 
because Pro might greatly affect the stereochemical structure. However, 
the inhibition by lysine alone was mitigated, and the degree of activation 
by threonine became large (not less than 120%). Thus the concerted 
inhibition was also desensitized. 
Thermal stability was investigated as an index of stability of the enzyme 
structure constructed by the introduction of mutation. The treatment 
condition was set at 55.degree. C. for 1.5 hour in which the activity of 
the wild type AK was about 80%. The Cys residue-introduced mutant strain, 
the Thr residue-introduced mutant strain, the Phe residue-introduced 
mutant strain, the Tyr residue-introduced mutant strain, and the Val 
residue-introduced mutant strain had higher stability than that of the 
wild type, among which the Val residue-introduced mutant strain had the 
highest stability (FIG. 6). 
Example 7 
Effect on L-Lysine Productivity by Introduction of Plasmids Containing 8 
Kinds of Mutant AK Genes and Wild Type AK Gene into Corynebacterium 
glutamicum Wild type strain 
Strains were prepared by introducing 8 kinds of the plasmids shown in Table 
3 into AJ12036 strain (FERM P-7559) as a wild type strain of 
Corynebacterium glutamicum (Brevibacterium lactofermentum) in the same 
manner as Example 3. The AK activity was measured for each of the strains. 
As shown in Table 5, the AK specific activities of the strains introduced 
with each of the plasmids were increased about 20-80 times as compared 
with the AK specific activity of the host. The desensitization degree of 
inhibition by lysine and/or threonine was similar to those in Example 6. 
The Pro residue-introduced mutant AK had the highest degree of 
desensitization of inhibition, in which the degree of desensitization of 
inhibition was greater than that of the Thr reside-introduced mutant AK, 
with respect to any of the inhibition by lysine alone, the inhibition by 
threonine alone, and the concerted inhibition by the both. 
TABLE 5 
______________________________________ 
Specific 
activity 
(mU/mg 
5 mM Lys 5 mM Thr 2 mM Lys 
protein) 
(%) (%) + Thr (%) 
______________________________________ 
AJ12036 5.6 52.0 82.0 7.0 
Wild type 316.4 52.7 86.8 6.2 
Thr 374.4 58.7 109.1 78.3 
Arg 197.4 41.4 106.8 58.6 
Cys 267.0 66.5 135.7 60.6 
Phe 447.7 14.6 105.0 32.4 
Pro 125.0 77.5 123.2 85.2 
Ser 406.8 55.0 114.4 37.0 
Tyr 425.6 16.1 104.8 32.2 
Val 448.9 60.5 103.5 75.5 
______________________________________ 
The lysine productivity was further evaluated by cultivation for the 9 
kinds of these strains by using a method similar to that in Example 3. The 
amount of produced lysine in each cultivation is as shown in Table 6. It 
is understood that the L-lysine productivity is remarkably improved by 
introducing the mutant AK plasmids. Especially, high accumulation of about 
25 g/l was exhibited in the mutant strains other than the Cys 
residue-introduced mutant strain and the Set residue-introduced mutant 
strain. The plasmid-holding ratio upon completion of the cultivation was 
approximately 100%, exhibiting high stability of the plasmids. 
TABLE 6 
______________________________________ 
Lys--HCl (g/l) 
Plasmid holding ratio (%) 
______________________________________ 
Wild type 0.00 100 
Thr 24.25 100 
Arg 24.56 100 
Cys 13.41 100 
Phe 25.14 100 
Pro 25.11 100 
Ser 5.72 100 
Tyr 25.12 100 
Val 25.02 100 
______________________________________ 
Industrial Applicability 
In the AK gene of Brevibacterium lactofermentum, Ala located at the 
position 279 in accordance with the amino acid numbers in SEQ ID NO: 3 or 
at the position 30 in accordance with the amino acid numbers in SEQ ID 
NO:5 was changed to amino acids other than acidic amino acids. Thus AK was 
obtained in which inhibition by threonine was completely desensitized, and 
consequently concerted inhibition by lysine+threonine was desensitized. 
Especially, AK was obtained, in which inhibition by lysine alone was 
partially desensitized, by changing the amino acid residue at the 
aforementioned position to Pro. AK having improved thermal stability was 
obtained by changing the aforementioned site to Val, Tyr, or Phe. The 
productivity of L-lysine could be remarkably increased by increasing the 
activity of such mutant AK in cells of coryneform bacteria. 
__________________________________________________________________________ 
SEQUENCE LISTING 
(1) GENERAL INFORMATION: 
(iii) NUMBER OF SEQUENCES: 24 
(2) INFORMATION FOR SEQ ID NO:1: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 1643 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: double 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: genomic DNA 
(iii) HYPOTHETICAL: NO 
(iv) ANTI-SENSE: NO 
(vi) ORIGINAL SOURCE: 
(A) ORGANISM: Corynebacterium glutamicum 
(B) STRAIN: ATCC 13869 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:1: 
TCGCGAAGTAGCACCTGTCACTTTTGTCTCAAATATTAAATCGAATATCAATATACGGTC60 
TGTTTATTGGAACGCATCCCAGTGGCTGAGACGCATCCGCTAAAGCCCCAGGAACCCTGT120 
GCAGAAAGAAAACACTCCTCTGGCTAGGTAGACACAGTTTATAAAGGTAGAGTTGAGCGG180 
GTAACTGTCAGCACGTAGATCGAAAGGTGCACAAAGGTGGCCCTGGTCGTACAGAAATAT240 
GGCGGTTCCTCGCTTGAGAGTGCGGAACGCATTAGAAACGTCGCTGAACGGATCGTTGCC300 
ACCAAGAAGGCTGGAAATGATGTCGTGGTTGTCTGCTCCGCAATGGGAGACACCACGGAT360 
GAACTTCTAGAACTTGCAGCGGCAGTGAATCCCGTTCCGCCAGCTCGTGAAATGGATATG420 
CTCCTGACTGCTGGTGAGCGTATTTCTAACGCTCTCGTCGCCATGGCTATTGAGTCCCTT480 
GGCGCAGAAGCTCAATCTTTCACTGGCTCTCAGGCTGGTGTGCTCACCACCGAGCGCCAC540 
GGAAACGCACGCATTGTTGACGTCACACCGGGTCGTGTGCGTGAAGCACTCGATGAGGGC600 
AAGATCTGCATTGTTGCTGGTTTTCAGGGTGTTAATAAAGAAACCCGCGATGTCACCACG660 
TTGGGTCGTGGTGGTTCTGACACCACTGCAGTTGCGTTGGCAGCTGCTTTGAACGCTGAT720 
GTGTGTGAGATTTACTCGGACGTTGACGGTGTGTATACCGCTGACCCGCGCATCGTTCCT780 
AATGCACAGAAGCTGGAAAAGCTCAGCTTCGAAGAAATGCTGGAACTTGCTGCTGTTGGC840 
TCCAAGATTTTGGTGCTGCGCAGTGTTGAATACGCTCGTGCATTCAATGTGCCACTTCGC900 
GTACGCTCGTCTTATAGTAATGATCCCGGCACTTTGATTGCCGGCTCTATGGAGGATATT960 
CCTGTGGAAGAAGCAGTCCTTACCGGTGTCGCAACCGACAAGTCCGAAGCCAAAGTAACC1020 
GTTCTGGGTATTTCCGATAAGCCAGGCGAGGCTGCCAAGGTTTTCCGTGCGTTGGCTGAT1080 
GCAGAAATCAACATTGACATGGTTCTGCAGAACGTCTCCTCTGTGGAAGACGGCACCACC1140 
GACATCACGTTCACCTGCCCTCGCGCTGACGGACGCCGTGCGATGGAGATCTTGAAGAAG1200 
CTTCAGGTTCAGGGCAACTGGACCAATGTGCTTTACGACGACCAGGTCGGCAAAGTCTCC1260 
CTCGTGGGTGCTGGCATGAAGTCTCACCCAGGTGTTACCGCAGAGTTCATGGAAGCTCTG1320 
CGCGATGTCAACGTGAACATCGAATTGATTTCCACCTCTGAGATCCGCATTTCCGTGCTG1380 
ATCCGTGAAGATGATCTGGATGCTGCTGCACGTGCATTGCATGAGCAGTTCCAGCTGGGC1440 
GGCGAAGACGAAGCCGTCGTTTATGCAGGCACCGGACGCTAAAGTTTTAAAGGAGTAGTT1500 
TTACAATGACCACCATCGCAGTTGTTGGTGCAACCGGCCAGGTCGGCCAGGTTATGCGCA1560 
CCCTTTTGGAAGAGCGCAATTTCCCAGCTGACACTGTTCGTTTCTTTGCTTCCCCGCGTT1620 
CCGCAGGCCGTAAGATTGAATTC1643 
(2) INFORMATION FOR SEQ ID NO:2: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 1643 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: double 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: genomic DNA 
(iii) HYPOTHETICAL: NO 
(iv) ANTI-SENSE: NO 
(vi) ORIGINAL SOURCE: 
(A) ORGANISM: Corynebacterium glutamicum 
(B) STRAIN: AJ3463 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:2: 
TCGCGAAGTAGCACCTGTCACTTTTGTCTCAAATATTAAATCGAATATCAATATACGGTC60 
TGTTTATTGGAACGCATCCCAGTGGCTGAGACGCATCCGCTAAAGCCCCAGGAACCCTGT120 
GCAGAAAGAAAACACTCCTCTGGCTAGGTAGACACAGTTTATAAAGGTAGAGTTGAGCGG180 
GTAACTGTCAGCACGTAGATCGAAAGGTGCACAAAGGTGGCCCTGGTCGTACAGAAATAT240 
GGCGGTTCCTCGCTTGAGAGTGCGGAACGCATTAGAAACGTCGCTGAACGGATCGTTGCC300 
ACCAAGAAGGCTGGAAATGATGTCGTGGTTGTCTGCTCCGCAATGGGAGACACCACGGAT360 
GAACTTCTAGAACTTGCAGCGGCAGTGAATCCCGTTCCGCCAGCTCGTGAAATGGATATG420 
CTCCTGACTGCTGGTGAGCGTATTTCTAACGCTCTCGTCGCCATGGCTATTGAGTCCCTT480 
GGCGCAGAAGCTCAATCTTTCACTGGCTCTCAGGCTGGTGTGCTCACCACCGAGCGCCAC540 
GGAAACGCACGCATTGTTGACGTCACACCGGGTCGTGTGCGTGAAGCACTCGATGAGGGC600 
AAGATCTGCATTGTTGCTGGTTTTCAGGGTGTTAATAAAGAAACCCGCGATGTCACCACG660 
TTGGGTCGTGGTGGTTCTGACACCACTGCAGTTGCGTTGGCAGCTGCTTTGAACGCTGAT720 
GTGTGTGAGATTTACTCGGACGTTGACGGTGTGTATACCGCTGACCCGCGCATCGTTCCT780 
AATGCACAGAAGCTGGAAAAGCTCAGCTTCGAAGAAATGCTGGAACTTGCTGCTGTTGGC840 
TCCAAGATTTTGGTGCTGCGCAGTGTTGAATACGCTCGTGCATTCAATGTGCCACTTCGC900 
GTACGCTCGTCTTATAGTAATGATCCCGGCACTTTGATTGCCGGCTCTATGGAGGATATT960 
CCTGTGGAAGAAGCAGTCCTTACCGGTGTCGCAACCGACAAGTCCGAAGCCAAAGTAACC1020 
GTTCTGGGTATTTCCGATAAGCCAGGCGAGACTGCCAAGGTTTTCCGTGCGTTGGCTGAT1080 
GCAGAAATCAACATTGACATGGTTCTGCAGAACGTCTCCTCTGTGGAAGACGGCACCACC1140 
GACATCACGTTCACCTGCCCTCGCGCTGACGGACGCCGTGCGATGGAGATCTTGAAGAAG1200 
CTTCAGGTTCAGGGCAACTGGACCAATGTGCTTTACGACGACCAGGTCGGCAAAGTCTCC1260 
CTCGTGGGTGCTGGCATGAAGTCTCACCCAGGTGTTACCGCAGAGTTCATGGAAGCTCTG1320 
CGCGATGTCAACGTGAACATCGAATTGATTTCCACCTCTGAGATCCGCATTTCCGTGCTG1380 
ATCCGTGAAGATGATCTGGATGCTGCTGCACGTGCATTGCATGAGCAGTTCCAGCTGGGC1440 
GGCGAAGACGAAGCCGTCGTTTATGCAGGCACCGGACGCTAAAGTTTTAAAGGAGTAGTT1500 
TTACAATGACCACCATCGCAGTTGTTGGTGCAACCGGCCAGGTCGGCCAGGTTATGCGCA1560 
CCCTTTTGGAAGAGCGCAATTTCCCAGCTGACACTGTTCGTTTCTTTGCTTCCCCGCGTT1620 
CCGCAGGCCGTAAGATTGAATTC1643 
(2) INFORMATION FOR SEQ ID NO:3: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 421 amino acids 
(B) TYPE: amino acid 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: peptide 
(iii) HYPOTHETICAL: NO 
(iv) ANTI-SENSE: NO 
(vi) ORIGINAL SOURCE: 
(A) ORGANISM: Corynebacterium glutamicum 
(B) STRAIN: ATCC13869 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:3: 
MetAlaLeuValValGlnLysTyrGlyGlySerSerLeuGluSerAla 
151015 
GluArgIleArgAsnValAlaGluArgIleValAlaThrLysLysAla 
202530 
GlyAsnAspValValValValCysSerAlaMetGlyAspThrThrAsp 
354045 
GluLeuLeuGluLeuAlaAlaAlaValAsnProValProProAlaArg 
505560 
GluMetAspMetLeuLeuThrAlaGlyGluArgIleSerAsnAlaLeu 
65707580 
ValAlaMetAlaIleGluSerLeuGlyAlaGluAlaGlnSerPheThr 
859095 
GlySerGlnAlaGlyValLeuThrThrGluArgHisGlyAsnAlaArg 
100105110 
IleValAspValThrProGlyArgValArgGluAlaLeuAspGluGly 
115120125 
LysIleCysIleValAlaGlyPheGlnGlyValAsnLysGluThrArg 
130135140 
AspValThrThrLeuGlyArgGlyGlySerAspThrThrAlaValAla 
145150155160 
LeuAlaAlaAlaLeuAsnAlaAspValCysGluIleTyrSerAspVal 
165170175 
AspGlyValTyrThrAlaAspProArgIleValProAsnAlaGlnLys 
180185190 
LeuGluLysLeuSerPheGluGluMetLeuGluLeuAlaAlaValGly 
195200205 
SerLysIleLeuValLeuArgSerValGluTyrAlaArgAlaPheAsn 
210215220 
ValProLeuArgValArgSerSerTyrSerAsnAspProGlyThrLeu 
225230235240 
IleAlaGlySerMetGluAspIleProValGluGluAlaValLeuThr 
245250255 
GlyValAlaThrAspLysSerGluAlaLysValThrValLeuGlyIle 
260265270 
SerAspLysProGlyGluAlaAlaLysValPheArgAlaLeuAlaAsp 
275280285 
AlaGluIleAsnIleAspMetValLeuGlnAsnValSerSerValGlu 
290295300 
AspGlyThrThrAspIleThrPheThrCysProArgAlaAspGlyArg 
305310315320 
ArgAlaMetGluIleLeuLysLysLeuGlnValGlnGlyAsnTrpThr 
325330335 
AsnValLeuTyrAspAspGlnValGlyLysValSerLeuValGlyAla 
340345350 
GlyMetLysSerHisProGlyValThrAlaGluPheMetGluAlaLeu 
355360365 
ArgAspValAsnValAsnIleGluLeuIleSerThrSerGluIleArg 
370375380 
IleSerValLeuIleArgGluAspAspLeuAspAlaAlaAlaArgAla 
385390395400 
LeuHisGluGlnPheGlnLeuGlyGlyGluAspGluAlaValValTyr 
405410415 
AlaGlyThrGlyArg 
420 
(2) INFORMATION FOR SEQ ID NO:4: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 421 amino acids 
(B) TYPE: amino acid 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: peptide 
(iii) HYPOTHETICAL: NO 
(iv) ANTI-SENSE: NO 
(vi) ORIGINAL SOURCE: 
(A) ORGANISM: Corynebacterium glutamicum 
(B) STRAIN: AJ3463 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:4: 
MetAlaLeuValValGlnLysTyrGlyGlySerSerLeuGluSerAla 
151015 
GluArgIleArgAsnValAlaGluArgIleValAlaThrLysLysAla 
202530 
GlyAsnAspValValValValCysSerAlaMetGlyAspThrThrAsp 
354045 
GluLeuLeuGluLeuAlaAlaAlaValAsnProValProProAlaArg 
505560 
GluMetAspMetLeuLeuThrAlaGlyGluArgIleSerAsnAlaLeu 
65707580 
ValAlaMetAlaIleGluSerLeuGlyAlaGluAlaGlnSerPheThr 
859095 
GlySerGlnAlaGlyValLeuThrThrGluArgHisGlyAsnAlaArg 
100105110 
IleValAspValThrProGlyArgValArgGluAlaLeuAspGluGly 
115120125 
LysIleCysIleValAlaGlyPheGlnGlyValAsnLysGluThrArg 
130135140 
AspValThrThrLeuGlyArgGlyGlySerAspThrThrAlaValAla 
145150155160 
LeuAlaAlaAlaLeuAsnAlaAspValCysGluIleTyrSerAspVal 
165170175 
AspGlyValTyrThrAlaAspProArgIleValProAsnAlaGlnLys 
180185190 
LeuGluLysLeuSerPheGluGluMetLeuGluLeuAlaAlaValGly 
195200205 
SerLysIleLeuValLeuArgSerValGluTyrAlaArgAlaPheAsn 
210215220 
ValProLeuArgValArgSerSerTyrSerAsnAspProGlyThrLeu 
225230235240 
IleAlaGlySerMetGluAspIleProValGluGluAlaValLeuThr 
245250255 
GlyValAlaThrAspLysSerGluAlaLysValThrValLeuGlyIle 
260265270 
SerAspLysProGlyGluThrAlaLysValPheArgAlaLeuAlaAsp 
275280285 
AlaGluIleAsnIleAspMetValLeuGlnAsnValSerSerValGlu 
290295300 
AspGlyThrThrAspIleThrPheThrCysProArgAlaAspGlyArg 
305310315320 
ArgAlaMetGluIleLeuLysLysLeuGlnValGlnGlyAsnTrpThr 
325330335 
AsnValLeuTyrAspAspGlnValGlyLysValSerLeuValGlyAla 
340345350 
GlyMetLysSerHisProGlyValThrAlaGluPheMetGluAlaLeu 
355360365 
ArgAspValAsnValAsnIleGluLeuIleSerThrSerGluIleArg 
370375380 
IleSerValLeuIleArgGluAspAspLeuAspAlaAlaAlaArgAla 
385390395400 
LeuHisGluGlnPheGlnLeuGlyGlyGluAspGluAlaValValTyr 
405410415 
AlaGlyThrGlyArg 
420 
(2) INFORMATION FOR SEQ ID NO:5: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 172 amino acids 
(B) TYPE: amino acid 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: peptide 
(iii) HYPOTHETICAL: NO 
(iv) ANTI-SENSE: NO 
(vi) ORIGINAL SOURCE: 
(A) ORGANISM: Corynebacterium glutamicum 
(B) STRAIN: ATCC13869 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:5: 
MetGluGluAlaValLeuThrGlyValAlaThrAspLysSerGluAla 
151015 
LysValThrValLeuGlyIleSerAspLysProGlyGluAlaAlaLys 
202530 
ValPheArgAlaLeuAlaAspAlaGluIleAsnIleAspMetValLeu 
354045 
GlnAsnValSerSerValGluAspGlyThrThrAspIleThrPheThr 
505560 
CysProArgAlaAspGlyArgArgAlaMetGluIleLeuLysLysLeu 
65707580 
GlnValGlnGlyAsnTrpThrAsnValLeuTyrAspAspGlnValGly 
859095 
LysValSerLeuValGlyAlaGlyMetLysSerHisProGlyValThr 
100105110 
AlaGluPheMetGluAlaLeuArgAspValAsnValAsnIleGluLeu 
115120125 
IleSerThrSerGluIleArgIleSerValLeuIleArgGluAspAsp 
130135140 
LeuAspAlaAlaAlaArgAlaLeuHisGluGlnPheGlnLeuGlyGly 
145150155160 
GluAspGluAlaValValTyrAlaGlyThrGlyArg 
165170 
(2) INFORMATION FOR SEQ ID NO:6: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 172 amino acids 
(B) TYPE: amino acid 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: peptide 
(iii) HYPOTHETICAL: NO 
(iv) ANTI-SENSE: NO 
(vi) ORIGINAL SOURCE: 
(A) ORGANISM: Corynebacterium glutamicum 
(B) STRAIN: AJ3463 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:6: 
MetGluGluAlaValLeuThrGlyValAlaThrAspLysSerGluAla 
151015 
LysValThrValLeuGlyIleSerAspLysProGlyGluThrAlaLys 
202530 
ValPheArgAlaLeuAlaAspAlaGluIleAsnIleAspMetValLeu 
354045 
GlnAsnValSerSerValGluAspGlyThrThrAspIleThrPheThr 
505560 
CysProArgAlaAspGlyArgArgAlaMetGluIleLeuLysLysLeu 
65707580 
GlnValGlnGlyAsnTrpThrAsnValLeuTyrAspAspGlnValGly 
859095 
LysValSerLeuValGlyAlaGlyMetLysSerHisProGlyValThr 
100105110 
AlaGluPheMetGluAlaLeuArgAspValAsnValAsnIleGluLeu 
115120125 
IleSerThrSerGluIleArgIleSerValLeuIleArgGluAspAsp 
130135140 
LeuAspAlaAlaAlaArgAlaLeuHisGluGlnPheGlnLeuGlyGly 
145150155160 
GluAspGluAlaValValTyrAlaGlyThrGlyArg 
165170 
(2) INFORMATION FOR SEQ ID NO:7: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 1643 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: double 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: genomic DNA 
(iii) HYPOTHETICAL: NO 
(iv) ANTI-SENSE: NO 
(vi) ORIGINAL SOURCE: 
(A) ORGANISM: Corynebacterium glutamicum 
(B) STRAIN: ATCC13869 
(ix) FEATURE: 
(A) NAME/KEY: mat peptide 
(B) LOCATION: 217..1482 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:7: 
TCGCGAAGTAGCACCTGTCACTTTTGTCTCAAATATTAAATCGAATATCAATATACGGTC60 
TGTTTATTGGAACGCATCCCAGTGGCTGAGACGCATCCGCTAAAGCCCCAGGAACCCTGT120 
GCAGAAAGAAAACACTCCTCTGGCTAGGTAGACACAGTTTATAAAGGTAGAGTTGAGCGG180 
GTAACTGTCAGCACGTAGATCGAAAGGTGCACAAAGGTGGCCCTGGTCGTACAG234 
MetAlaLeuValValGln 
15 
AAATATGGCGGTTCCTCGCTTGAGAGTGCGGAACGCATTAGAAACGTC282 
LysTyrGlyGlySerSerLeuGluSerAlaGluArgIleArgAsnVal 
101520 
GCTGAACGGATCGTTGCCACCAAGAAGGCTGGAAATGATGTCGTGGTT330 
AlaGluArgIleValAlaThrLysLysAlaGlyAsnAspValValVal 
253035 
GTCTGCTCCGCAATGGGAGACACCACGGATGAACTTCTAGAACTTGCA378 
ValCysSerAlaMetGlyAspThrThrAspGluLeuLeuGluLeuAla 
404550 
GCGGCAGTGAATCCCGTTCCGCCAGCTCGTGAAATGGATATGCTCCTG426 
AlaAlaValAsnProValProProAlaArgGluMetAspMetLeuLeu 
55606570 
ACTGCTGGTGAGCGTATTTCTAACGCTCTCGTCGCCATGGCTATTGAG474 
ThrAlaGlyGluArgIleSerAsnAlaLeuValAlaMetAlaIleGlu 
758085 
TCCCTTGGCGCAGAAGCTCAATCTTTCACTGGCTCTCAGGCTGGTGTG522 
SerLeuGlyAlaGluAlaGlnSerPheThrGlySerGlnAlaGlyVal 
9095100 
CTCACCACCGAGCGCCACGGAAACGCACGCATTGTTGACGTCACACCG570 
LeuThrThrGluArgHisGlyAsnAlaArgIleValAspValThrPro 
105110115 
GGTCGTGTGCGTGAAGCACTCGATGAGGGCAAGATCTGCATTGTTGCT618 
GlyArgValArgGluAlaLeuAspGluGlyLysIleCysIleValAla 
120125130 
GGTTTTCAGGGTGTTAATAAAGAAACCCGCGATGTCACCACGTTGGGT666 
GlyPheGlnGlyValAsnLysGluThrArgAspValThrThrLeuGly 
135140145150 
CGTGGTGGTTCTGACACCACTGCAGTTGCGTTGGCAGCTGCTTTGAAC714 
ArgGlyGlySerAspThrThrAlaValAlaLeuAlaAlaAlaLeuAsn 
155160165 
GCTGATGTGTGTGAGATTTACTCGGACGTTGACGGTGTGTATACCGCT762 
AlaAspValCysGluIleTyrSerAspValAspGlyValTyrThrAla 
170175180 
GACCCGCGCATCGTTCCTAATGCACAGAAGCTGGAAAAGCTCAGCTTC810 
AspProArgIleValProAsnAlaGlnLysLeuGluLysLeuSerPhe 
185190195 
GAAGAAATGCTGGAACTTGCTGCTGTTGGCTCCAAGATTTTGGTGCTG858 
GluGluMetLeuGluLeuAlaAlaValGlySerLysIleLeuValLeu 
200205210 
CGCAGTGTTGAATACGCTCGTGCATTCAATGTGCCACTTCGCGTACGC906 
ArgSerValGluTyrAlaArgAlaPheAsnValProLeuArgValArg 
215220225230 
TCGTCTTATAGTAATGATCCCGGCACTTTGATTGCCGGCTCTATGGAG954 
SerSerTyrSerAsnAspProGlyThrLeuIleAlaGlySerMetGlu 
235240245 
GATATTCCTGTGGAAGAAGCAGTCCTTACCGGTGTCGCAACCGACAAG1002 
AspIleProValGluGluAlaValLeuThrGlyValAlaThrAspLys 
250255260 
TCCGAAGCCAAAGTAACCGTTCTGGGTATTTCCGATAAGCCAGGCGAG1050 
SerGluAlaLysValThrValLeuGlyIleSerAspLysProGlyGlu 
265270275 
GCTGCCAAGGTTTTCCGTGCGTTGGCTGATGCAGAAATCAACATTGAC1098 
AlaAlaLysValPheArgAlaLeuAlaAspAlaGluIleAsnIleAsp 
280285290 
ATGGTTCTGCAGAACGTCTCCTCTGTGGAAGACGGCACCACCGACATC1146 
MetValLeuGlnAsnValSerSerValGluAspGlyThrThrAspIle 
295300305310 
ACGTTCACCTGCCCTCGCGCTGACGGACGCCGTGCGATGGAGATCTTG1194 
ThrPheThrCysProArgAlaAspGlyArgArgAlaMetGluIleLeu 
315320325 
AAGAAGCTTCAGGTTCAGGGCAACTGGACCAATGTGCTTTACGACGAC1242 
LysLysLeuGlnValGlnGlyAsnTrpThrAsnValLeuTyrAspAsp 
330335340 
CAGGTCGGCAAAGTCTCCCTCGTGGGTGCTGGCATGAAGTCTCACCCA1290 
GlnValGlyLysValSerLeuValGlyAlaGlyMetLysSerHisPro 
345350355 
GGTGTTACCGCAGAGTTCATGGAAGCTCTGCGCGATGTCAACGTGAAC1338 
GlyValThrAlaGluPheMetGluAlaLeuArgAspValAsnValAsn 
360365370 
ATCGAATTGATTTCCACCTCTGAGATCCGCATTTCCGTGCTGATCCGT1386 
IleGluLeuIleSerThrSerGluIleArgIleSerValLeuIleArg 
375380385390 
GAAGATGATCTGGATGCTGCTGCACGTGCATTGCATGAGCAGTTCCAG1434 
GluAspAspLeuAspAlaAlaAlaArgAlaLeuHisGluGlnPheGln 
395400405 
CTGGGCGGCGAAGACGAAGCCGTCGTTTATGCAGGCACCGGACGCTAA1482 
LeuGlyGlyGluAspGluAlaValValTyrAlaGlyThrGlyArg 
410415420421 
AGTTTTAAAGGAGTAGTTTTACAATGACCACCATCGCAGTTGTTGGTGCAACCGGCCAGG1542 
TCGGCCAGGTTATGCGCACCCTTTTGGAAGAGCGCAATTTCCCAGCTGACACTGTTCGTT1602 
TCTTTGCTTCCCCGCGTTCCGCAGGCCGTAAGATTGAATTC1643 
(2) INFORMATION FOR SEQ ID NO:8: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 1643 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: double 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: genomic DNA 
(iii) HYPOTHETICAL: NO 
(iv) ANTI-SENSE: NO 
(vi) ORIGINAL SOURCE: 
(A) ORGANISM: Corynebacterium glutamicum 
(B) STRAIN: AJ3463 
(ix) FEATURE: 
(A) NAME/KEY: mat peptide 
(B) LOCATION: 217..1482 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:8: 
TCGCGAAGTAGCACCTGTCACTTTTGTCTCAAATATTAAATCGAATATCAATATACGGTC60 
TGTTTATTGGAACGCATCCCAGTGGCTGAGACGCATCCGCTAAAGCCCCAGGAACCCTGT120 
GCAGAAAGAAAACACTCCTCTGGCTAGGTAGACACAGTTTATAAAGGTAGAGTTGAGCGG180 
GTAACTGTCAGCACGTAGATCGAAAGGTGCACAAAGGTGGCCCTGGTCGTACAG234 
MetAlaLeuValValGln 
15 
AAATATGGCGGTTCCTCGCTTGAGAGTGCGGAACGCATTAGAAACGTC282 
LysTyrGlyGlySerSerLeuGluSerAlaGluArgIleArgAsnVal 
101520 
GCTGAACGGATCGTTGCCACCAAGAAGGCTGGAAATGATGTCGTGGTT330 
AlaGluArgIleValAlaThrLysLysAlaGlyAsnAspValValVal 
253035 
GTCTGCTCCGCAATGGGAGACACCACGGATGAACTTCTAGAACTTGCA378 
ValCysSerAlaMetGlyAspThrThrAspGluLeuLeuGluLeuAla 
404550 
GCGGCAGTGAATCCCGTTCCGCCAGCTCGTGAAATGGATATGCTCCTG426 
AlaAlaValAsnProValProProAlaArgGluMetAspMetLeuLeu 
55606570 
ACTGCTGGTGAGCGTATTTCTAACGCTCTCGTCGCCATGGCTATTGAG474 
ThrAlaGlyGluArgIleSerAsnAlaLeuValAlaMetAlaIleGlu 
758085 
TCCCTTGGCGCAGAAGCTCAATCTTTCACTGGCTCTCAGGCTGGTGTG522 
SerLeuGlyAlaGluAlaGlnSerPheThrGlySerGlnAlaGlyVal 
9095100 
CTCACCACCGAGCGCCACGGAAACGCACGCATTGTTGACGTCACACCG570 
LeuThrThrGluArgHisGlyAsnAlaArgIleValAspValThrPro 
105110115 
GGTCGTGTGCGTGAAGCACTCGATGAGGGCAAGATCTGCATTGTTGCT618 
GlyArgValArgGluAlaLeuAspGluGlyLysIleCysIleValAla 
120125130 
GGTTTTCAGGGTGTTAATAAAGAAACCCGCGATGTCACCACGTTGGGT666 
GlyPheGlnGlyValAsnLysGluThrArgAspValThrThrLeuGly 
135140145150 
CGTGGTGGTTCTGACACCACTGCAGTTGCGTTGGCAGCTGCTTTGAAC714 
ArgGlyGlySerAspThrThrAlaValAlaLeuAlaAlaAlaLeuAsn 
155160165 
GCTGATGTGTGTGAGATTTACTCGGACGTTGACGGTGTGTATACCGCT762 
AlaAspValCysGluIleTyrSerAspValAspGlyValTyrThrAla 
170175180 
GACCCGCGCATCGTTCCTAATGCACAGAAGCTGGAAAAGCTCAGCTTC810 
AspProArgIleValProAsnAlaGlnLysLeuGluLysLeuSerPhe 
185190195 
GAAGAAATGCTGGAACTTGCTGCTGTTGGCTCCAAGATTTTGGTGCTG858 
GluGluMetLeuGluLeuAlaAlaValGlySerLysIleLeuValLeu 
200205210 
CGCAGTGTTGAATACGCTCGTGCATTCAATGTGCCACTTCGCGTACGC906 
ArgSerValGluTyrAlaArgAlaPheAsnValProLeuArgValArg 
215220225230 
TCGTCTTATAGTAATGATCCCGGCACTTTGATTGCCGGCTCTATGGAG954 
SerSerTyrSerAsnAspProGlyThrLeuIleAlaGlySerMetGlu 
235240245 
GATATTCCTGTGGAAGAAGCAGTCCTTACCGGTGTCGCAACCGACAAG1002 
AspIleProValGluGluAlaValLeuThrGlyValAlaThrAspLys 
250255260 
TCCGAAGCCAAAGTAACCGTTCTGGGTATTTCCGATAAGCCAGGCGAG1050 
SerGluAlaLysValThrValLeuGlyIleSerAspLysProGlyGlu 
265270275 
ACTGCCAAGGTTTTCCGTGCGTTGGCTGATGCAGAAATCAACATTGAC1098 
ThrAlaLysValPheArgAlaLeuAlaAspAlaGluIleAsnIleAsp 
280285290 
ATGGTTCTGCAGAACGTCTCCTCTGTGGAAGACGGCACCACCGACATC1146 
MetValLeuGlnAsnValSerSerValGluAspGlyThrThrAspIle 
295300305310 
ACGTTCACCTGCCCTCGCGCTGACGGACGCCGTGCGATGGAGATCTTG1194 
ThrPheThrCysProArgAlaAspGlyArgArgAlaMetGluIleLeu 
315320325 
AAGAAGCTTCAGGTTCAGGGCAACTGGACCAATGTGCTTTACGACGAC1242 
LysLysLeuGlnValGlnGlyAsnTrpThrAsnValLeuTyrAspAsp 
330335340 
CAGGTCGGCAAAGTCTCCCTCGTGGGTGCTGGCATGAAGTCTCACCCA1290 
GlnValGlyLysValSerLeuValGlyAlaGlyMetLysSerHisPro 
345350355 
GGTGTTACCGCAGAGTTCATGGAAGCTCTGCGCGATGTCAACGTGAAC1338 
GlyValThrAlaGluPheMetGluAlaLeuArgAspValAsnValAsn 
360365370 
ATCGAATTGATTTCCACCTCTGAGATCCGCATTTCCGTGCTGATCCGT1386 
IleGluLeuIleSerThrSerGluIleArgIleSerValLeuIleArg 
375380385390 
GAAGATGATCTGGATGCTGCTGCACGTGCATTGCATGAGCAGTTCCAG1434 
GluAspAspLeuAspAlaAlaAlaArgAlaLeuHisGluGlnPheGln 
395400405 
CTGGGCGGCGAAGACGAAGCCGTCGTTTATGCAGGCACCGGACGCTAA1482 
LeuGlyGlyGluAspGluAlaValValTyrAlaGlyThrGlyArg 
410415420421 
AGTTTTAAAGGAGTAGTTTTACAATGACCACCATCGCAGTTGTTGGTGCAACCGGCCAGG1542 
TCGGCCAGGTTATGCGCACCCTTTTGGAAGAGCGCAATTTCCCAGCTGACACTGTTCGTT1602 
TCTTTGCTTCCCCGCGTTCCGCAGGCCGTAAGATTGAATTC1643 
(2) INFORMATION FOR SEQ ID NO:9: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 1643 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: double 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: genomic DNA 
(iii) HYPOTHETICAL: NO 
(iv) ANTI-SENSE: NO 
(vi) ORIGINAL SOURCE: 
(A) ORGANISM: Corynebacterium glutamicum 
(B) STRAIN: ATCC13869 
(ix) FEATURE: 
(A) NAME/KEY: mat peptide 
(B) LOCATION: 964..1482 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:9: 
TCGCGAAGTAGCACCTGTCACTTTTGTCTCAAATATTAAATCGAATATCAATATACGGTC60 
TGTTTATTGGAACGCATCCCAGTGGCTGAGACGCATCCGCTAAAGCCCCAGGAACCCTGT120 
GCAGAAAGAAAACACTCCTCTGGCTAGGTAGACACAGTTTATAAAGGTAGAGTTGAGCGG180 
GTAACTGTCAGCACGTAGATCGAAAGGTGCACAAAGGTGGCCCTGGTCGTACAGAAATAT240 
GGCGGTTCCTCGCTTGAGAGTGCGGAACGCATTAGAAACGTCGCTGAACGGATCGTTGCC300 
ACCAAGAAGGCTGGAAATGATGTCGTGGTTGTCTGCTCCGCAATGGGAGACACCACGGAT360 
GAACTTCTAGAACTTGCAGCGGCAGTGAATCCCGTTCCGCCAGCTCGTGAAATGGATATG420 
CTCCTGACTGCTGGTGAGCGTATTTCTAACGCTCTCGTCGCCATGGCTATTGAGTCCCTT480 
GGCGCAGAAGCTCAATCTTTCACTGGCTCTCAGGCTGGTGTGCTCACCACCGAGCGCCAC540 
GGAAACGCACGCATTGTTGACGTCACACCGGGTCGTGTGCGTGAAGCACTCGATGAGGGC600 
AAGATCTGCATTGTTGCTGGTTTTCAGGGTGTTAATAAAGAAACCCGCGATGTCACCACG660 
TTGGGTCGTGGTGGTTCTGACACCACTGCAGTTGCGTTGGCAGCTGCTTTGAACGCTGAT720 
GTGTGTGAGATTTACTCGGACGTTGACGGTGTGTATACCGCTGACCCGCGCATCGTTCCT780 
AATGCACAGAAGCTGGAAAAGCTCAGCTTCGAAGAAATGCTGGAACTTGCTGCTGTTGGC840 
TCCAAGATTTTGGTGCTGCGCAGTGTTGAATACGCTCGTGCATTCAATGTGCCACTTCGC900 
GTACGCTCGTCTTATAGTAATGATCCCGGCACTTTGATTGCCGGCTCTATGGAGGATATT960 
CCTGTGGAAGAAGCAGTCCTTACCGGTGTCGCAACCGACAAGTCCGAA1008 
MetGluGluAlaValLeuThrGlyValAlaThrAspLysSerGlu 
151015 
GCCAAAGTAACCGTTCTGGGTATTTCCGATAAGCCAGGCGAGGCTGCC1056 
AlaLysValThrValLeuGlyIleSerAspLysProGlyGluAlaAla 
202530 
AAGGTTTTCCGTGCGTTGGCTGATGCAGAAATCAACATTGACATGGTT1104 
LysValPheArgAlaLeuAlaAspAlaGluIleAsnIleAspMetVal 
354045 
CTGCAGAACGTCTCCTCTGTGGAAGACGGCACCACCGACATCACGTTC1152 
LeuGlnAsnValSerSerValGluAspGlyThrThrAspIleThrPhe 
505560 
ACCTGCCCTCGCGCTGACGGACGCCGTGCGATGGAGATCTTGAAGAAG1200 
ThrCysProArgAlaAspGlyArgArgAlaMetGluIleLeuLysLys 
657075 
CTTCAGGTTCAGGGCAACTGGACCAATGTGCTTTACGACGACCAGGTC1248 
LeuGlnValGlnGlyAsnTrpThrAsnValLeuTyrAspAspGlnVal 
80859095 
GGCAAAGTCTCCCTCGTGGGTGCTGGCATGAAGTCTCACCCAGGTGTT1296 
GlyLysValSerLeuValGlyAlaGlyMetLysSerHisProGlyVal 
100105110 
ACCGCAGAGTTCATGGAAGCTCTGCGCGATGTCAACGTGAACATCGAA1344 
ThrAlaGluPheMetGluAlaLeuArgAspValAsnValAsnIleGlu 
115120125 
TTGATTTCCACCTCTGAGATCCGCATTTCCGTGCTGATCCGTGAAGAT1392 
LeuIleSerThrSerGluIleArgIleSerValLeuIleArgGluAsp 
130135140 
GATCTGGATGCTGCTGCACGTGCATTGCATGAGCAGTTCCAGCTGGGC1440 
AspLeuAspAlaAlaAlaArgAlaLeuHisGluGlnPheGlnLeuGly 
145150155 
GGCGAAGACGAAGCCGTCGTTTATGCAGGCACCGGACGCTAAAGTTTTAA1490 
GlyGluAspGluAlaValValTyrAlaGlyThrGlyArg 
160165170172 
AGGAGTAGTTTTACAATGACCACCATCGCAGTTGTTGGTGCAACCGGCCAGGTCGGCCAG1550 
GTTATGCGCACCCTTTTGGAAGAGCGCAATTTCCCAGCTGACACTGTTCGTTTCTTTGCT1610 
TCCCCGCGTTCCGCAGGCCGTAAGATTGAATTC1643 
(2) INFORMATION FOR SEQ ID NO:10: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 1643 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: double 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: genomic DNA 
(iii) HYPOTHETICAL: NO 
(iv) ANTI-SENSE: NO 
(vi) ORIGINAL SOURCE: 
(A) ORGANISM: Corynebacterium glutamicum 
(B) STRAIN: AJ3463 
(ix) FEATURE: 
(A) NAME/KEY: mat peptide 
(B) LOCATION: 964..1482 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:10: 
TCGCGAAGTAGCACCTGTCACTTTTGTCTCAAATATTAAATCGAATATCAATATACGGTC60 
TGTTTATTGGAACGCATCCCAGTGGCTGAGACGCATCCGCTAAAGCCCCAGGAACCCTGT120 
GCAGAAAGAAAACACTCCTCTGGCTAGGTAGACACAGTTTATAAAGGTAGAGTTGAGCGG180 
GTAACTGTCAGCACGTAGATCGAAAGGTGCACAAAGGTGGCCCTGGTCGTACAGAAATAT240 
GGCGGTTCCTCGCTTGAGAGTGCGGAACGCATTAGAAACGTCGCTGAACGGATCGTTGCC300 
ACCAAGAAGGCTGGAAATGATGTCGTGGTTGTCTGCTCCGCAATGGGAGACACCACGGAT360 
GAACTTCTAGAACTTGCAGCGGCAGTGAATCCCGTTCCGCCAGCTCGTGAAATGGATATG420 
CTCCTGACTGCTGGTGAGCGTATTTCTAACGCTCTCGTCGCCATGGCTATTGAGTCCCTT480 
GGCGCAGAAGCTCAATCTTTCACTGGCTCTCAGGCTGGTGTGCTCACCACCGAGCGCCAC540 
GGAAACGCACGCATTGTTGACGTCACACCGGGTCGTGTGCGTGAAGCACTCGATGAGGGC600 
AAGATCTGCATTGTTGCTGGTTTTCAGGGTGTTAATAAAGAAACCCGCGATGTCACCACG660 
TTGGGTCGTGGTGGTTCTGACACCACTGCAGTTGCGTTGGCAGCTGCTTTGAACGCTGAT720 
GTGTGTGAGATTTACTCGGACGTTGACGGTGTGTATACCGCTGACCCGCGCATCGTTCCT780 
AATGCACAGAAGCTGGAAAAGCTCAGCTTCGAAGAAATGCTGGAACTTGCTGCTGTTGGC840 
TCCAAGATTTTGGTGCTGCGCAGTGTTGAATACGCTCGTGCATTCAATGTGCCACTTCGC900 
GTACGCTCGTCTTATAGTAATGATCCCGGCACTTTGATTGCCGGCTCTATGGAGGATATT960 
CCTGTGGAAGAAGCAGTCCTTACCGGTGTCGCAACCGACAAGTCCGAA1008 
ValGluGluAlaValLeuThrGlyValAlaThrAspLysSerGlu 
151015 
GCCAAAGTAACCGTTCTGGGTATTTCCGATAAGCCAGGCGAGACTGCC1056 
AlaLysValThrValLeuGlyIleSerAspLysProGlyGluThrAla 
202530 
AAGGTTTTCCGTGCGTTGGCTGATGCAGAAATCAACATTGACATGGTT1104 
LysValPheArgAlaLeuAlaAspAlaGluIleAsnIleAspMetVal 
354045 
CTGCAGAACGTCTCCTCTGTGGAAGACGGCACCACCGACATCACGTTC1152 
LeuGlnAsnValSerSerValGluAspGlyThrThrAspIleThrPhe 
505560 
ACCTGCCCTCGCGCTGACGGACGCCGTGCGATGGAGATCTTGAAGAAG1200 
ThrCysProArgAlaAspGlyArgArgAlaMetGluIleLeuLysLys 
657075 
CTTCAGGTTCAGGGCAACTGGACCAATGTGCTTTACGACGACCAGGTC1248 
LeuGlnValGlnGlyAsnTrpThrAsnValLeuTyrAspAspGlnVal 
80859095 
GGCAAAGTCTCCCTCGTGGGTGCTGGCATGAAGTCTCACCCAGGTGTT1296 
GlyLysValSerLeuValGlyAlaGlyMetLysSerHisProGlyVal 
100105110 
ACCGCAGAGTTCATGGAAGCTCTGCGCGATGTCAACGTGAACATCGAA1344 
ThrAlaGluPheMetGluAlaLeuArgAspValAsnValAsnIleGlu 
115120125 
TTGATTTCCACCTCTGAGATCCGCATTTCCGTGCTGATCCGTGAAGAT1392 
LeuIleSerThrSerGluIleArgIleSerValLeuIleArgGluAsp 
130135140 
GATCTGGATGCTGCTGCACGTGCATTGCATGAGCAGTTCCAGCTGGGC1440 
AspLeuAspAlaAlaAlaArgAlaLeuHisGluGlnPheGlnLeuGly 
145150155 
GGCGAAGACGAAGCCGTCGTTTATGCAGGCACCGGACGCTAAAGTTTTAA1490 
GlyGluAspGluAlaValValTyrAlaGlyThrGlyArg 
160165170172 
AGGAGTAGTTTTACAATGACCACCATCGCAGTTGTTGGTGCAACCGGCCAGGTCGGCCAG1550 
GTTATGCGCACCCTTTTGGAAGAGCGCAATTTCCCAGCTGACACTGTTCGTTTCTTTGCT1610 
TCCCCGCGTTCCGCAGGCCGTAAGATTGAATTC1643 
(2) INFORMATION FOR SEQ ID NO:11: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 1263 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: double 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: genomic DNA 
(iii) HYPOTHETICAL: NO 
(iv) ANTI-SENSE: NO 
(vi) ORIGINAL SOURCE: 
(A) ORGANISM: Corynebacterium glutamicum 
(B) STRAIN: ATCC13869 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:11: 
GTGGCCCTGGTCGTACAGAAATATGGCGGTTCCTCGCTTGAGAGTGCGGAACGCATTAGA60 
AACGTCGCTGAACGGATCGTTGCCACCAAGAAGGCTGGAAATGATGTCGTGGTTGTCTGC120 
TCCGCAATGGGAGACACCACGGATGAACTTCTAGAACTTGCAGCGGCAGTGAATCCCGTT180 
CCGCCAGCTCGTGAAATGGATATGCTCCTGACTGCTGGTGAGCGTATTTCTAACGCTCTC240 
GTCGCCATGGCTATTGAGTCCCTTGGCGCAGAAGCTCAATCTTTCACTGGCTCTCAGGCT300 
GGTGTGCTCACCACCGAGCGCCACGGAAACGCACGCATTGTTGACGTCACACCGGGTCGT360 
GTGCGTGAAGCACTCGATGAGGGCAAGATCTGCATTGTTGCTGGTTTTCAGGGTGTTAAT420 
AAAGAAACCCGCGATGTCACCACGTTGGGTCGTGGTGGTTCTGACACCACTGCAGTTGCG480 
TTGGCAGCTGCTTTGAACGCTGATGTGTGTGAGATTTACTCGGACGTTGACGGTGTGTAT540 
ACCGCTGACCCGCGCATCGTTCCTAATGCACAGAAGCTGGAAAAGCTCAGCTTCGAAGAA600 
ATGCTGGAACTTGCTGCTGTTGGCTCCAAGATTTTGGTGCTGCGCAGTGTTGAATACGCT660 
CGTGCATTCAATGTGCCACTTCGCGTACGCTCGTCTTATAGTAATGATCCCGGCACTTTG720 
ATTGCCGGCTCTATGGAGGATATTCCTGTGGAAGAAGCAGTCCTTACCGGTGTCGCAACC780 
GACAAGTCCGAAGCCAAAGTAACCGTTCTGGGTATTTCCGATAAGCCAGGCGAGGCTGCC840 
AAGGTTTTCCGTGCGTTGGCTGATGCAGAAATCAACATTGACATGGTTCTGCAGAACGTC900 
TCCTCTGTGGAAGACGGCACCACCGACATCACGTTCACCTGCCCTCGCGCTGACGGACGC960 
CGTGCGATGGAGATCTTGAAGAAGCTTCAGGTTCAGGGCAACTGGACCAATGTGCTTTAC1020 
GACGACCAGGTCGGCAAAGTCTCCCTCGTGGGTGCTGGCATGAAGTCTCACCCAGGTGTT1080 
ACCGCAGAGTTCATGGAAGCTCTGCGCGATGTCAACGTGAACATCGAATTGATTTCCACC1140 
TCTGAGATCCGCATTTCCGTGCTGATCCGTGAAGATGATCTGGATGCTGCTGCACGTGCA1200 
TTGCATGAGCAGTTCCAGCTGGGCGGCGAAGACGAAGCCGTCGTTTATGCAGGCACCGGA1260 
CGC1263 
(2) INFORMATION FOR SEQ ID NO:12: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 1263 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: double 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: genomic DNA 
(iii) HYPOTHETICAL: NO 
(iv) ANTI-SENSE: NO 
(vi) ORIGINAL SOURCE: 
(A) ORGANISM: Corynebacterium glutamicum 
(B) STRAIN: AJ3463 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:12: 
GTGGCCCTGGTCGTACAGAAATATGGCGGTTCCTCGCTTGAGAGTGCGGAACGCATTAGA60 
AACGTCGCTGAACGGATCGTTGCCACCAAGAAGGCTGGAAATGATGTCGTGGTTGTCTGC120 
TCCGCAATGGGAGACACCACGGATGAACTTCTAGAACTTGCAGCGGCAGTGAATCCCGTT180 
CCGCCAGCTCGTGAAATGGATATGCTCCTGACTGCTGGTGAGCGTATTTCTAACGCTCTC240 
GTCGCCATGGCTATTGAGTCCCTTGGCGCAGAAGCTCAATCTTTCACTGGCTCTCAGGCT300 
GGTGTGCTCACCACCGAGCGCCACGGAAACGCACGCATTGTTGACGTCACACCGGGTCGT360 
GTGCGTGAAGCACTCGATGAGGGCAAGATCTGCATTGTTGCTGGTTTTCAGGGTGTTAAT420 
AAAGAAACCCGCGATGTCACCACGTTGGGTCGTGGTGGTTCTGACACCACTGCAGTTGCG480 
TTGGCAGCTGCTTTGAACGCTGATGTGTGTGAGATTTACTCGGACGTTGACGGTGTGTAT540 
ACCGCTGACCCGCGCATCGTTCCTAATGCACAGAAGCTGGAAAAGCTCAGCTTCGAAGAA600 
ATGCTGGAACTTGCTGCTGTTGGCTCCAAGATTTTGGTGCTGCGCAGTGTTGAATACGCT660 
CGTGCATTCAATGTGCCACTTCGCGTACGCTCGTCTTATAGTAATGATCCCGGCACTTTG720 
ATTGCCGGCTCTATGGAGGATATTCCTGTGGAAGAAGCAGTCCTTACCGGTGTCGCAACC780 
GACAAGTCCGAAGCCAAAGTAACCGTTCTGGGTATTTCCGATAAGCCAGGCGAGACTGCC840 
AAGGTTTTCCGTGCGTTGGCTGATGCAGAAATCAACATTGACATGGTTCTGCAGAACGTC900 
TCCTCTGTGGAAGACGGCACCACCGACATCACGTTCACCTGCCCTCGCGCTGACGGACGC960 
CGTGCGATGGAGATCTTGAAGAAGCTTCAGGTTCAGGGCAACTGGACCAATGTGCTTTAC1020 
GACGACCAGGTCGGCAAAGTCTCCCTCGTGGGTGCTGGCATGAAGTCTCACCCAGGTGTT1080 
ACCGCAGAGTTCATGGAAGCTCTGCGCGATGTCAACGTGAACATCGAATTGATTTCCACC1140 
TCTGAGATCCGCATTTCCGTGCTGATCCGTGAAGATGATCTGGATGCTGCTGCACGTGCA1200 
TTGCATGAGCAGTTCCAGCTGGGCGGCGAAGACGAAGCCGTCGTTTATGCAGGCACCGGA1260 
CGC1263 
(2) INFORMATION FOR SEQ ID NO:13: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 516 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: double 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: genomic DNA 
(iii) HYPOTHETICAL: NO 
(iv) ANTI-SENSE: NO 
(vi) ORIGINAL SOURCE: 
(A) ORGANISM: Corynebacterium glutamicum 
(B) STRAIN: ATCC13869 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:13: 
GTGGAAGAAGCAGTCCTTACCGGTGTCGCAACCGACAAGTCCGAAGCCAAAGTAACCGTT60 
CTGGGTATTTCCGATAAGCCAGGCGAGGCTGCCAAGGTTTTCCGTGCGTTGGCTGATGCA120 
GAAATCAACATTGACATGGTTCTGCAGAACGTCTCCTCTGTGGAAGACGGCACCACCGAC180 
ATCACGTTCACCTGCCCTCGCGCTGACGGACGCCGTGCGATGGAGATCTTGAAGAAGCTT240 
CAGGTTCAGGGCAACTGGACCAATGTGCTTTACGACGACCAGGTCGGCAAAGTCTCCCTC300 
GTGGGTGCTGGCATGAAGTCTCACCCAGGTGTTACCGCAGAGTTCATGGAAGCTCTGCGC360 
GATGTCAACGTGAACATCGAATTGATTTCCACCTCTGAGATCCGCATTTCCGTGCTGATC420 
CGTGAAGATGATCTGGATGCTGCTGCACGTGCATTGCATGAGCAGTTCCAGCTGGGCGGC480 
GAAGACGAAGCCGTCGTTTATGCAGGCACCGGACGC516 
(2) INFORMATION FOR SEQ ID NO:14: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 516 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: double 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: genomic DNA 
(iii) HYPOTHETICAL: NO 
(iv) ANTI-SENSE: NO 
(vi) ORIGINAL SOURCE: 
(A) ORGANISM: Corynebacterium glutamicum 
(B) STRAIN: AJ3463 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:14: 
GTGGAAGAAGCAGTCCTTACCGGTGTCGCAACCGACAAGTCCGAAGCCAAAGTAACCGTT60 
CTGGGTATTTCCGATAAGCCAGGCGAGACTGCCAAGGTTTTCCGTGCGTTGGCTGATGCA120 
GAAATCAACATTGACATGGTTCTGCAGAACGTCTCCTCTGTGGAAGACGGCACCACCGAC180 
ATCACGTTCACCTGCCCTCGCGCTGACGGACGCCGTGCGATGGAGATCTTGAAGAAGCTT240 
CAGGTTCAGGGCAACTGGACCAATGTGCTTTACGACGACCAGGTCGGCAAAGTCTCCCTC300 
GTGGGTGCTGGCATGAAGTCTCACCCAGGTGTTACCGCAGAGTTCATGGAAGCTCTGCGC360 
GATGTCAACGTGAACATCGAATTGATTTCCACCTCTGAGATCCGCATTTCCGTGCTGATC420 
CGTGAAGATGATCTGGATGCTGCTGCACGTGCATTGCATGAGCAGTTCCAGCTGGGCGGC480 
GAAGACGAAGCCGTCGTTTATGCAGGCACCGGACGC516 
(2) INFORMATION FOR SEQ ID NO:15: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 23 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: single 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: synthetic DNA 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:15: 
TCGCGAAGTAGCACCTGTCACTT23 
(2) INFORMATION FOR SEQ ID NO:16: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 21 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: single 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: synthetic DNA 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:16: 
ACGGAATTCAATCTTACGGCC21 
(2) INFORMATION FOR SEQ ID NO:17: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 23 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: single 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: synthetic DNA 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:17: 
GCCAGGCGAGCGTGCCAAGGTTT23 
(2) INFORMATION FOR SEQ ID NO:18: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 23 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: single 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: synthetic DNA 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:18: 
GCCAGGCGAGGATGCCAAGGTTT23 
(2) INFORMATION FOR SEQ ID NO:19: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 23 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: single 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: synthetic DNA 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:19: 
GCCAGGCGAGTGTGCCAAGGTTT23 
(2) INFORMATION FOR SEQ ID NO:20: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 23 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: single 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: synthetic DNA 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:20: 
GCCAGGCGAGTTTGCCAAGGTTT23 
(2) INFORMATION FOR SEQ ID NO:21: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 23 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: single 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: synthetic DNA 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:21: 
GCCAGGCGAGCCTGCCAAGGTTT23 
(2) INFORMATION FOR SEQ ID NO:22: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 23 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: single 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: synthetic DNA 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:22: 
GCCAGGCGAGTCTGCCAAGGTTT23 
(2) INFORMATION FOR SEQ ID NO:23: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 23 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: single 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: synthetic DNA 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:23: 
GCCAGGCGAGTATGCCAAGGTTT23 
(2) INFORMATION FOR SEQ ID NO:24: 
(i) SEQUENCE CHARACTERISTICS: 
(A) LENGTH: 23 nucleotides 
(B) TYPE: nucleic acid 
(C) STRANDEDNESS: single 
(D) TOPOLOGY: linear 
(ii) MOLECULE TYPE: synthetic DNA 
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:24: 
GCCAGGCGAGGTTGCCAAGGTTT23 
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