bluuebunny/crossref_metadata_2025_split huggingface dataset:
An Oligoribonuclease Gene in
Streptomyces griseus
Authors: Ohnishi Yasuo , Nishiyama Yoko , Sato Rie , Kameyama Shogo , Horinouchi Sueharu 
Published: July 2002

ABSTRACT

In
Streptomyces griseus
, A-factor (2-isocapryloyl-3
R
-hydroxymethyl-γ-butyrolactone) serves as a microbial hormone that switches on many genes required for streptomycin production and morphological development. An open reading frame (Orf1) showing high sequence similarity to oligoribonucleases of various origins is present just downstream of
adpA
, one of the A-factor-dependent genes. Orf1 was named OrnA (oligoribonuclease A) because it showed 3′-to-5′ exo-oligoribonuclease activity, releasing [
32
P]CMP from ApCpC[
32
P]pC used as a substrate. Reverse transcription-PCR and S1 nuclease mapping analyses revealed that
ornA
was transcribed from two promoters; one was a developmentally regulated, A-factor-dependent promoter in front of
adpA
, and the other was a constitutive promoter in front of the
ornA
coding sequence. Transcription of
ornA
was thus additively enhanced at the initiation stage for secondary metabolism and aerial mycelium formation.
ornA
-disrupted strains grew slowly and scarcely formed aerial mycelium.
ornA
homologues were distributed in a wide variety of
Streptomyces
species, including
S. coelicolor
A3(2), as determined by Southern hybridization analysis. Disruption of the
ornA
homologue in
S. coelicolor
A3(2) also caused phenotypes similar to those of the
S. griseus ΔornA
strains. The OrnA oligoribonucleases in
Streptomyces
species are therefore not essential but play an important role in vegetative growth and in the initiation of differentiation.
Cloning and Characterization of the Glucooligosaccharide Catabolic Pathway β-Glucan Glucohydrolase and Cellobiose Phosphorylase in the Marine HyperthermophileThermotoga neapolitana
Authors: Yernool Dinesh A. , McCarthy James K. , Eveleigh Douglas E. , Bok Jin-Duck 
Published: July 2002

ABSTRACTCharacterization inThermotoga neapolitanaof a catabolic gene cluster encoding two glycosyl hydrolases, 1,4-β-d-glucan glucohydrolase (GghA) and cellobiose phosphorylase (CbpA), and the apparent absence of a cellobiohydrolase (Cbh) suggest a nonconventional pathway for glucan utilization inThermotogales. GghA purified fromT. neapolitanais a 52.5-kDa family 1 glycosyl hydrolase with optimal activity at pH 6.5 and 95°C. GghA releases glucose from soluble glucooligomers, with a preference for longer oligomers:kcat/Kmvalues are 155.2, 76.0, and 9.9 mM−1s−1for cellotetraose, cellotriose, and cellobiose, respectively. GghA has broad substrate specificity, with specific activities of 236 U/mg towards cellobiose and 251 U/mg towards lactose. Withp-nitrophenyl-β-glucoside as the substrate, GghA exhibits biphasic kinetic behavior, involving both substrate- and end product-directed activation. Its capacity for transglycosylation is a factor in this activation. Cloning ofgghArevealed a contiguous upstream gene (cbpA) encoding a 93.5-kDa cellobiose phosphorylase. Recombinant CbpA has optimal activity at pH 5.0 and 85°C. It has specific activity of 11.8 U/mg and aKmof 1.42 mM for cellobiose, but shows no activity towards other disaccharides or cellotriose. With its single substrate specificity and lowKmfor cellobiose (compared to GghA'sKmof 28.6 mM), CbpA may be the primary enzyme for attacking cellobiose inThermotogaspp. By phosphorolysis of cellobiose, CbpA releases one activated glucosyl molecule while conserving one ATP molecule per disaccharide. CbpA is the first hyperthermophilic cellobiose phosphorylase to be characterized.
The NosX and NirX Proteins of
Paracoccus denitrificans
Are Functional Homologues: Their Role in Maturation of Nitrous Oxide Reductase
Authors: Saunders Neil F. W. , Hornberg Jorrit J. , Reijnders Willem N. M. , Westerhoff Hans V. , de Vries Simon , van Spanning Rob J. M. 
Published: July 2002

ABSTRACT

The
nos
(nitrous oxide reductase) operon of
Paracoccus denitrificans
contains a
nosX
gene homologous to those found in the
nos
operons of other denitrifiers. NosX is also homologous to NirX, which is so far unique to
P. denitrificans
. Single mutations of these genes did not result in any apparent phenotype, but a double
nosX nirX
mutant was unable to reduce nitrous oxide. Promoter-
lacZ
assays and immunoblotting against nitrous oxide reductase showed that the defect was not due to failure of expression of
nosZ
, the structural gene for nitrous oxide reductase. Electron paramagnetic resonance spectroscopy showed that nitrous oxide reductase in cells of the double mutant lacked the Cu
A
center. A twin-arginine motif in both NosX and NirX suggests that the NosX proteins are exported to the periplasm via the TAT translocon.
Characterization of
ssfR
and
ssgA
, Two Genes Involved in Sporulation of
Streptomyces griseus
Authors: Jiang Hao , Kendrick Kathleen E. 
Published: July 2002

ABSTRACT

In the presence of cefoxitin, which inhibits septum formation during sporulation,
Streptomyces griseus
is unable to sporulate, retaining the sonication sensitivity of nonsporulating hyphae. Cefoxitin- and sonication-resistant mutant SKK2600 was isolated and showed many morphological differences from its parental strain. A 3.6-kb DNA fragment that complemented the mutations of SKK2600 contained two open reading frames (ORFs), either of which could complement SKK2600. One ORF, designated
ssfR
, encoded a protein containing a potential DNA-binding helix-turn-helix motif close to its N terminus. SsfR is similar to members of a large family of transcriptional regulators, particularly IclR of
Escherichia coli
. The second ORF was identified as
ssgA
, a previously described sporulation gene from
S. griseus
(S. Kawamoto and J. C. Ensign, Actinomycetology 9:136–151, 1995). A point mutation of C to T seven nucleotides upstream of the UGA stop codon of
ssfR
was responsible for the phenotype of isolated mutant strain SKK2600. Surprisingly, this mutation should not change the primary structure of SsfR. The
ssfR
and
ssgA
disruption mutants were constructed and showed the “white” mutant phenotype, with some growth medium dependence. In addition, the
ssfR
null mutant sporulated ectopically in phosphate starvation medium.
Isolation of Regulated Genes of the Cyanobacterium
Synechocystis
sp. Strain PCC 6803 by Differential Display
Authors: Bhaya Devaki , Vaulot Daniel , Amin Pinky , Takahashi Akiko Watanabe , Grossman Arthur R. 
Published: July 2002

ABSTRACT

Global identification of differentially regulated genes in prokaryotes is constrained because the mRNA does not have a 3′ polyadenylation extension; this precludes specific separation of mRNA from rRNA and tRNA and synthesis of cDNAs from the entire mRNA population. Knowledge of the entire genome sequence of
Synechocystis
sp. strain PCC 6803 has enabled us to develop a differential display procedure that takes advantage of a short palindromic sequence that is dispersed throughout the
Synechocystis
sp. strain PCC 6803 genome. This sequence, designated the HIP (highly iterated palindrome) element, occurs in approximately half of the
Synechocystis
sp. strain PCC 6803 genes but is absent in rRNA and tRNA genes. To determine the feasibility of exploiting the HIP element, alone or in combination with specific primer subsets, for analyzing differential gene expression, we used HIP-based primers to identify light intensity-regulated genes. Several gene fragments, including those encoding ribosomal proteins and phycobiliprotein subunits, were differentially amplified from RNA templates derived from cells grown in low light or exposed to high light for 3 h. One novel finding was that expression of certain genes of the
pho
regulon, which are under the control of environmental phosphate levels, were markedly elevated in high light. High-light activation of
pho
regulon genes correlated with elevated growth rates that occur when the cells are transferred from low to high light. These results suggest that in high light, the rate of growth of
Synechocystis
sp. strain PCC 6803 exceeds its capacity to assimilate phosphate, which, in turn, may trigger a phosphate starvation response and activation of the
pho
regulon.
Identification and Mapping of Sigma-54 Promoters in
Chlamydia trachomatis
Authors: Mathews Sarah A. , Timms Peter 
Published: July 2002

ABSTRACT

The first ς
54
promoters in
Chlamydia trachomatis
L2 were mapped upstream of hypothetical proteins CT652.1 and CT683. Comparative genomics indicated that these ς
54
promoters and potential upstream activation binding sites are conserved in orthologous
C. trachomatis
D,
C. trachomatis
mouse pneumonitis strain, and
Chlamydia pneumoniae
(CWL029 and AR39) genes.
Role of the Carboxy Terminus of
Escherichia coli
FtsA in Self-Interaction and Cell Division
Authors: Yim Lucía , Vandenbussche Guy , Mingorance Jesús , Rueda Sonsoles , Casanova Mercedes , Ruysschaert Jean-Marie , Vicente Miguel 
Published: July 2002

ABSTRACT

The role of the carboxy terminus of the
Escherichia coli
cell division protein FtsA in bacterial division has been studied by making a series of short sequential deletions spanning from residue 394 to 420. Deletions as short as 5 residues destroy the biological function of the protein. Residue W415 is essential for the localization of the protein into septal rings. Overexpression of the
ftsA
alleles harboring these deletions caused a coiled cell phenotype previously described for another carboxy-terminal mutation (Gayda et al., J. Bacteriol. 174:5362–5370, 1992), suggesting that an interaction of FtsA with itself might play a role in its function. The existence of such an interaction was demonstrated using the yeast two-hybrid system and a protein overlay assay. Even these short deletions are sufficient for impairing the interaction of the truncated FtsA forms with the wild-type protein in the yeast two-hybrid system. The existence of additional interactions between FtsA molecules, involving other domains, can be postulated from the interaction properties shown by the FtsA deletion mutant forms, because although unable to interact with the wild-type and with FtsAΔ1, they can interact with themselves and cross-interact with each other. The secondary structures of an extensive deletion, FtsAΔ27, and the wild-type protein are indistinguishable when analyzed by Fourier transform infrared spectroscopy, and moreover, FtsAΔ27 retains the ability to bind ATP. These results indicate that deletion of the carboxy-terminal 27 residues does not alter substantially the structure of the protein and suggest that the loss of biological function of the carboxy-terminal deletion mutants might be related to the modification of their interacting properties.
Complementary Metal Ion Specificity of the Metal-Citrate Transporters CitM and CitH of
Bacillus subtilis
Authors: Krom Bastiaan P. , Warner Jessica B. , Konings Wil N. , Lolkema Juke S. 
Published: July 2002

ABSTRACT

Citrate uptake in
Bacillus subtilis
is stimulated by a wide range of divalent metal ions. The metal ions were separated into two groups based on the expression pattern of the uptake system. The two groups correlated with the metal ion specificity of two homologous
B. subtilis
secondary citrate transporters, CitM and CitH, upon expression in
Escherichia coli
. CitM transported citrate in complex with Mg
2+
, Ni
2+
, Mn
2+
, Co
2+
, and Zn
2+
but not in complex with Ca
2+
, Ba
2+
, and Sr
2+
. CitH transported citrate in complex with Ca
2+
, Ba
2+
, and Sr
2+
but not in complex with Mg
2+
, Ni
2+
, Mn
2+
, Co
2+
, and Zn
2+
. Both transporters did not transport free citrate. Nevertheless, free citrate uptake could be demonstrated in
B. subtilis
, indicating the expression of at least a third citrate transporter, whose identity is not known. For both the CitM and CitH transporters it was demonstrated that the metal ion promoted citrate uptake and, vice versa, that citrate promoted uptake of the metal ion, indicating that the complex is the transported species. The results indicate that CitM and CitH are secondary transporters that transport complexes of divalent metal ions and citrate but with a complementary metal ion specificity. The potential physiological function of the two transporters is discussed.
Characterization of a Novel Outer Membrane Hemin-Binding Protein of
Porphyromonas gingivalis
Authors: Dashper S. G. , Hendtlass A. , Slakeski N. , Jackson C. , Cross K. J. , Brownfield L. , Hamilton R. , Barr I. , Reynolds E. C. 
Published: July 2002

ABSTRACT

Porphyromonas gingivalis
is a gram-negative, anaerobic coccobacillus that has been implicated as a major etiological agent in the development of chronic periodontitis. In this paper, we report the characterization of a protein, IhtB (iron heme transport; formerly designated Pga30), that is an outer membrane hemin-binding protein potentially involved in iron assimilation by
P. gingivalis
. IhtB was localized to the cell surface of
P. gingivalis
by Western blot analysis of a Sarkosyl-insoluble outer membrane preparation and by immunocytochemical staining of whole cells using IhtB peptide-specific antisera. The protein, released from the cell surface, was shown to bind to hemin using hemin-agarose. The growth of heme-limited, but not heme-replete,
P. gingivalis
cells was inhibited by preincubation with IhtB peptide-specific antisera. The
ihtB
gene was located between an open reading frame encoding a putative TonB-linked outer membrane receptor and three open reading frames that have sequence similarity to ATP binding cassette transport system operons in other bacteria. Analysis of the deduced amino acid sequence of IhtB showed significant similarity to the
Salmonella typhimurium
protein CbiK, a cobalt chelatase that is structurally related to the ATP-independent family of ferrochelatases. Molecular modeling indicated that the IhtB amino acid sequence could be threaded onto the CbiK fold with the IhtB structural model containing the active-site residues critical for chelatase activity. These results suggest that IhtB is a peripheral outer membrane chelatase that may remove iron from heme prior to uptake by
P. gingivalis
.
Evidence for Horizontal Gene Transfer in Evolution of Elongation Factor Tu in Enterococci
Authors: Ke Danbing , Boissinot Maurice , Huletsky Ann , Picard François J. , Frenette Johanne , Ouellette Marc , Roy Paul H. , Bergeron Michel G. 
Published: July 2002

ABSTRACTThe elongation factor Tu, encoded bytufgenes, is a GTP binding protein that plays a central role in protein synthesis. One to threetufgenes per genome are present, depending on the bacterial species. Most low-G+C-content gram-positive bacteria carry only onetufgene. We have designed degenerate PCR primers derived from consensus sequences of thetufgene to amplify partialtufsequences from 17 enterococcal species and other phylogenetically related species. The amplified DNA fragments were sequenced either by direct sequencing or by sequencing cloned inserts containing putative amplicons. Two differenttufgenes (tufAandtufB) were found in 11 enterococcal species, includingEnterococcus avium,Enterococcus casseliflavus,Enterococcus dispar,Enterococcus durans,Enterococcus faecium,Enterococcus gallinarum,Enterococcus hirae,Enterococcus malodoratus,Enterococcus mundtii,Enterococcus pseudoavium, andEnterococcus raffinosus. For the other six enterococcal species (Enterococcus cecorum,Enterococcus columbae,Enterococcus faecalis,Enterococcus sulfureus,Enterococcus saccharolyticus, andEnterococcus solitarius), only thetufAgene was present. Based on 16S rRNA gene sequence analysis, the 11 species having twotufgenes all have a common ancestor, while the six species having only one copy diverged from the enterococcal lineage before that common ancestor. The presence of one or two copies of thetufgene in enterococci was confirmed by Southern hybridization. Phylogenetic analysis oftufsequences demonstrated that the enterococcaltufAgene branches with theBacillus,Listeria, andStaphylococcusgenera, while the enterococcaltufBgene clusters with the generaStreptococcusandLactococcus. Primary structure analysis showed that four amino acid residues encoded within the sequenced regions are conserved and unique to the enterococcaltufBgenes and thetufgenes of streptococci andLactococcus lactis. The data suggest that an ancestral streptococcus or a streptococcus-related species may have horizontally transferred atufgene to the common ancestor of the 11 enterococcal species which now carry twotufgenes.
Molecular Evolution of Virulence in Natural Field Strains of
Xanthomonas campestris
pv. vesicatoria
Authors: Gassmann Walter , Dahlbeck Douglas , Chesnokova Olga , Minsavage Gerald V. , Jones Jeffrey B. , Staskawicz Brian J. 
Published: July 2002

ABSTRACT

The
avrBs2
avirulence gene of the bacterial plant pathogen
Xanthomonas campestris
pv. vesicatoria triggers disease resistance in pepper plants containing the
Bs2
resistance gene and contributes to bacterial virulence on susceptible host plants. We studied the effects of the pepper
Bs2
gene on the evolution of
avrBs2
by characterizing the molecular basis for virulence of 20
X. campestris
pv. vesicatoria field strains that were isolated from disease spots on previously resistant
Bs2
pepper plants. All field strains tested were complemented by a wild-type copy of
avrBs2
in their ability to trigger disease resistance on
Bs2
plants. DNA sequencing revealed four mutant alleles of
avrBs2
, two of which consisted of insertions or deletions of 5 nucleotides in a repetitive region of
avrBs2
. The other two
avrBs2
alleles were characterized by point mutations with resulting single amino acid changes (R403P or A410D). We generated isogenic
X. campestris
pv. vesicatoria strains by chromosomal
avrBs2
gene exchange to study the effects of these mutations on the dual functions of
avrBs2
in enhancing bacterial virulence and inducing plant resistance by in planta bacterial growth experiments. The deletion of 5 nucleotides led to loss of
avrBs2
-induced resistance on
Bs2
pepper plants and abolition of
avrBs2
-mediated enhancement of fitness on susceptible plants. Significantly, the point mutations led to minimal reduction in virulence function of
avrBs2
on susceptible pepper plants, with either minimal (R403P allele) or an intermediate level of (A410D allele) triggering of resistance on
Bs2
plants. Consistent with the divergent selection pressures on
avrBs2
exerted by the
Bs2
resistance gene, our results show that
avrBs2
is evolving to decrease detection by the
Bs2
gene while at the same time maintaining its virulence function.
Sensitive Genetic Screen for Protease Activity Based on a Cyclic AMP Signaling Cascade in
Escherichia coli
Authors: Dautin Nathalie , Karimova Gouzel , Ullmann Agnes , Ladant Daniel 
Published: July 2002

ABSTRACT

We describe a genetic system that allows in vivo screening or selection of site-specific proteases and of their cognate-specific inhibitors in
Escherichia coli
. This genetic test is based on the specific proteolysis of a signaling enzyme, the adenylate cyclase (AC) of
Bordetella pertussis
. As a model system we used the human immunodeficiency virus (HIV) protease. When an HIV protease processing site, p5, was inserted in frame into the AC polypeptide, the resulting ACp5 protein retained enzymatic activity and, when expressed in an
E. coli cya
strain, restored the Cya
+
phenotype. The HIV protease coexpressed in the same cells resulted in cleavage and inactivation of ACp5; the cells became Cya
−
. When the entire HIV protease, including its adjacent processing sites, was inserted into the AC polypeptide, the resulting AC-HIV-Pr fusion protein, expressed in
E. coli cya
, was autoproteolysed and inactivated: the cells displayed Cya
−
phenotype. In the presence of the protease inhibitor indinavir or saquinavir, AC-HIV-Pr autoproteolysis was inhibited and the AC activity of the fusion protein was preserved; the cells were Cya
+
. Protease variants resistant to particular inhibitors could be easily distinguished from the wild type, as the cells displayed a Cya
−
phenotype in the presence of these inhibitors. This genetic test could represent a powerful approach to screen for new proteolytic activities and for novel protease inhibitors. It could also be used to detect in patients undergoing highly active antiretroviral therapy the emergence of HIV variants harboring antiprotease-resistant proteases.
Transcription Activation by a Variety of AraC/XylS Family Activators Does Not Depend on the Class II-Specific Activation Determinant in the N-Terminal Domain of the RNA Polymerase Alpha Subunit
Authors: Egan Susan M. , Pease Andrew J. , Lang Jeffrey , Li Xiang , Rao Vydehi , Gillette William K. , Ruiz Raquel , Ramos Juan L. , Wolf Richard E. 
Published: July 2002

ABSTRACT
The N-terminal domain of the RNA polymerase α subunit (α-NTD) was tested for a role in transcription activation by a variety of AraC/XylS family members. Based on substitutions at residues 162 to 165 and an extensive genetic screen we conclude that α-NTD is not an activation target for these activators.
A Novel Operon Encoding Formaldehyde Fixation: the Ribulose Monophosphate Pathway in the Gram-Positive Facultative Methylotrophic Bacterium
Mycobacterium gastri
MB19
Authors: Mitsui Ryoji , Sakai Yasuyoshi , Yasueda Hisashi , Kato Nobuo 
Published: July 2002

ABSTRACT

A 4.2-kb
Pst
I fragment harboring the gene cluster of the ribulose monophosphate (RuMP) pathway for formaldehyde fixation was identified in the chromosome of a gram-positive, facultative methylotroph,
Mycobacterium gastri
MB19, by using the coding region of 3-hexulose-6-phosphate synthase (HPS) as the hybridization probe. The
Pst
I fragment contained three complete open reading frames (ORFs) which encoded from the 5′ end, a DNA-binding regulatory protein (
rmpR
), 6-phospho-3-hexuloisomerase (PHI;
rmpB
), and HPS (
rmpA
). Sequence analysis suggested that
rmpA
and
rmpB
constitute an operon, and Northern blot analysis of RNA extracted from bacteria grown under various conditions suggested that the expression of the two genes is similarly regulated at the transcriptional level. A similarity search revealed that the proteins encoded by
rmpA
and
rmpB
in
M. gastri
MB19 show high similarity to the unidentified proteins of nonmethylotrophic prokaryotes, including bacteria and anaerobic archaea. The clusters in the phylogenetic tree of the HPS protein of
M. gastri
MB19 and those in the phylogenetic tree of the PHI protein were nearly identical, which implies that these two formaldehyde-fixing genes evolved as a pair. These findings give new insight into the acquisition of the formaldehyde fixation pathway during the evolution of diverse microorganisms.
Requirement for the Enzymes Acetoacetyl Coenzyme A Synthetase and Poly-3-Hydroxybutyrate (PHB) Synthase for Growth of
Sinorhizobium meliloti
on PHB Cycle Intermediates
Authors: Cai Guo-qin , Driscoll Brian T. , Charles Trevor C. 
Published: July 2002

ABSTRACT

We have identified two
Sinorhizobium meliloti
chromosomal loci affecting the poly-3-hydroxybutyrate degradation pathway. One locus was identified as the gene
acsA
, encoding acetoacetyl coenzyme A (acetoacetyl-CoA) synthetase. Analysis of the
acsA
nucleotide sequence revealed that this gene encodes a putative protein with a molecular weight of 72,000 that shows similarity to acetyl-CoA synthetase in other organisms. Acetyl-CoA synthetase activity was not affected in cell extracts of glucose-grown
acsA
::Tn
5
mutants; instead, acetoacetyl-CoA synthetase activity was drastically reduced. These findings suggest that acetoacetyl-CoA synthetase, rather than CoA transferase, activates acetoacetate to acetoacetyl-CoA in the
S. meliloti
poly-3-hydroxybutyrate cycle. The second locus was identified as
phbC
, encoding poly-3-hydroxybutyrate synthase, and was found to be required for synthesis of poly-3-hydroxybutyrate deposits.