bluuebunny/crossref_metadata_2025 huggingface dataset:
Variation in Escherichia coli buoyant density measured in Percoll gradients
Authors: Woldringh C L , Binnerts J S , Mans A
Published: January 2020

Escherichia coli B/r cells, centrifuged to equilibrium in either self-generating or preformed gradients of Percoll, banded at an average density of 1.080 to 1.100 g/ml, depending on their growth rate and the temperature of centrifugation. Cells cultured in gradient material (70% Percoll) exhibited the same average density. At the various growth rates examined, the density of the individual cells in a steady-state population varied by less than 1% of the mean in E. coli strains B/r and B, as well as K-12. Electron microscope analysis of the density fractions of both fast- and slow-growing E. coli B/r populations suggested a small increase in density during cell constriction.
Multiple nutritional requirements of lactobacilli: genetic lesions affecting amino acid biosynthetic pathways
Authors: Morishita T , Deguchi Y , Yajima M , Sakurai T , Yura T
Published: January 2020

Genetic lesions responsible for amino acid requirements in several species of multiple auxotrophic lactobacilli were investigated. Systematic attempts were made to isolate mutants that could grow in the absence of each of the amino acids required by the parental strains of Lactobacillus plantarum, L. casei, L. helveticus, and L. acidophilus. After treatment with appropriate mutagens, such mutants could be obtained with respect to many but not all required amino acids. Successful isolation of mutants for a given amino acid means that a minor genetic lesion reparable by single-step mutations affects its biosynthesis; a failure to isolate mutants suggests the involvement of more extensive lesions. Analysis of these results as well as the specific requirements exhibited by the parental strains revealed certain regularities; some of the biosynthetic pathways for individual amino acids were virtually unaffected by more extensive lesions in at least species tested, whereas others were affected by more extensive lesions in at least some species. Both the number and the kind of pathways affected by extensive lesions differed appreciably among different species. Furthermore, the growth response of the parental strains to some putative amino acid precursors revealed a clear correlation between the extent of genetic lesions and the occurrence and location of a genetic block(s) for a given pathway. These findings are discussed in relation to the phylogeny, ecology, and evolution of lactic acid bacteria.
Inhibition of Bacillus cereus spore outgrowth by covalent modification of a sulfhydryl group by nitrosothiol and iodoacetate
Authors: Morris S L , Hansen J N
Published: January 2020

Nitrosothiols with the general structure RSN==O were studied as a model system of bacteriostatic action toward outgrowing bacterial spores. With a Taft plot analysis, the influence of the structure of the R group on the inhibitory effectiveness of a series of nitrosothiols showed that effectiveness as an inhibitor of Bacillus cereus T outgrowth correlated with the electron withdrawal of R, but that size, shape, charge, hydrophobicity, and transportability had little influence. This was interpreted to mean that nitrosothiols do not traverse the membrane to act. The Taft plot together with competition data between nitrosothiol and iodoacetate indicated that the mode of nitrosothiol action is the covalent modification of a sulfhydryl group, probably to form RSN(OH)--SX, where --SX is derived from a sensitive spore sulfhydryl group. Cooperativity effects indicated that outgrowth inhibition is accompanied by a conformational change occurring upon sulfhydryl group modification, which is communicated among at least three to five subunits. Uptake of label during spore germination indicated that most of the sulfhydryl groups which can be modified are associated with the inhibitory event. These data suggest that this sulfhydryl group may be sufficiently unique that inhibitors designed to interfere specifically with it could have value as bacteriostatic agents.
Cell wall modification resulting from in vitro induction of L-phase variants of Nocardia asteroides
Authors: Beaman B L , Bourgeois A L , Moring S E
Published: January 2020

The chemical composition of the cell walls of several L-form revertants derived from Nocardia asteroides 10905 was determined at different stages of growth. It was observed that each L-form revertant had a cell well that differed from that of the parental strain when grown under identical conditions. In some strains the peptidolipid and mycolic acid components were affected the most, whereas in other strains the fatty acid, sugar, and mycolic acid moieties were altered. Shifts in mycolic acid size were prominent, whereas the basic peptidoglycan structure appeared to be affected the least. Both the method used to induce the L-form of N. asteroides 10905 and the length of time these organisms were maintained in the wall-less state affected the degree of cell wall modification during the reversion process. Thus, removal of the cell wall appeared to potentiate and select for mutational alterations within the cell envelope of N. asteroides, and these changes resulted in altered cellular and colonial morphology.
Genetic mapping of a linked cluster of ribosomal ribonucleic acid genes in Bacillus subtilis
Authors: Wilson F E , Hoch J A , Bott K
Published: January 2020

A ribosomal ribonucleic acid gene set consisting of genes for 16S, 23S, 5S, and 4S ribonucleic acid species has been genetically mapped to a position between the markers recG13 and abrB74 on the Bacillus subtilis chromosome and designated rrnA. A ribosomal mutation, ksgA, was found to be linked to rrnA. This places rrnA in a region of the chromosome where ribosome-related genes occur but that is not directly adjacent to the major cluster of ribosome-related markers.
Dominant spore color mutants of Aspergillus nidulans defective in germination and sexual development
Authors: Kurtz M B , Champe S P
Published: January 2020

The ascomycete Aspergillus nidulans produces green conidia (asexual spores). Recessive mutants which produce yellow conidia have been previously isolated from haploid strains and have been shown to be deficient in laccase (diphenol oxidase), an enzyme that requires copper for activity. Using a diploid parent strain, we isolated dominant yellow conidial mutants which, in the haploid state, produced even less laccase activity than a recessive mutant. Three isolates of such mutants behaved similarly and define a single complementation group (yB) on chromosome VIII distinct from the yA locus on chromosome I defined by recessive mutants. Unlike yA mutants, whose only discernable phenotype is their conidial color, yB mutants are pleiotropic: conidial germination was delayed relative to the wild type, and sexual development was blocked at an early stage. The three phenotypes of yB mutants were expressed on yeast extract-glucose medium containing 1.6 microM of added copper. When copper was added to above 5 microM, all three phenotypes were remediated, and near wild-type levels of laccase were produced. We conclude that yB mutants have a reduced availability of copper. The dominance of yB mutants could result, for example, from an alteration in transport or storage of copper. Using an immunological assay, we detected no laccase antigenic cross-reacting material in yB mutants grown on medium of low copper content. We conclude that either the synthesis or the stability of laccase is copper dependent.
Regulation of glutamate dehydrogenase in Bacillus subtilis
Authors: Kane J F , Wakim J , Fischer R S
Published: January 2020

The activity of the nicotinamide adenine dinucleotide-dependent glutamate dehydrogenase in Bacillus subtilis was influenced by the carbon source, but not the nitrogen source, in the growth medium. The highest specific activity for this enzyme was found when B. subtilis was grown in a minimal or rich medium that contained glutamate as the carbon source. It is proposed that glutamate dehydrogenase serves a catabolic function in the metabolism of glutamate, is induced by glutamate, and is subject to catabolite repression.
Defective specialized SP beta transducing bacteriophages of Bacillus subtilis that carry the sup-3 or sup-44 gene
Authors: Lipsky R H , Rosenthal R , Zahler S A
Published: January 2020

We isolated defective specialized transducing phages of SP beta that carry one of the extracistronic suppressors, sup-3 or sup-44. Lysates containing these phages can be used in a simple spot test to determine whether an auxotrophic mutation can be suppressed. The sup-3 and sup-44 mutations are distinct, in that their suppression patterns differ for the markers hisA1, metC3, and thr-5; and they are not alleles.
Homology between the invertible deoxyribonucleic acid sequence that controls flagellar-phase variation in Salmonella sp. and deoxyribonucleic acid sequences in other organisms
Authors: Szekely E , Simon M
Published: January 2020

The invertible deoxyribonucleic acid (DNA) segment cloned from Salmonella sp. was radioactively labeled and used as a probe to search for homologous sequences by Southern hybridization. Only one copy of the invertible segment could be found on the Salmonella sp. genome. Partial sequence homology with the invertible region was detected in bacteriophage Mu and P1 DNA by low-stringency hybridization. Under these conditions, no homology was detected with Escherichia coli DNA. A strain of Salmonella sp. defective in phase variation carrying the vH2- allele was also analyzed by DNA-DNA hybridization. The results show that there is sequence divergence between diphasic and vH2- strains within the invertible sequence.
Sexual agglutination in Saccharomyces cerevisiae
Authors: Terrance K , Lipke P N
Published: January 2020

Treatment of either mating type of Saccharomyces cerevisiae with the appropriate sex pheromone increased cell-cell binding in a modified cocentrifugation assay. Constitutive agglutination of haploids was qualitatively similar to pheromone-induced agglutination. Regardless of exposure to pheromone, agglutinable combinations of cells exhibited maximal binding across similar ranges of ionic strength, pH, and temperature. Binding of all combinations was inhibited by 8 M urea, 1 M pyridine, or 0.05% sodium dodecyl sulfate. From alpha-cells we solubilized and partially purified an inhibitor of a-cell agglutinability. This inhibitor reversibly masked all a-cell adhesion sites and inactivated pheromone-treated and control cells with similar kinetics. The inhibitor behaved as a homogeneous species in heat inactivation experiments. Based on these results, we proposed a model for pheromone effects on agglutination in S. cerevisiae.
Purification and Some Properties of Carbon Monoxide Dehydrogenase from
Pseudomonas carboxydohydrogena
Authors: Kim Y. M. , Hegeman G. D.
Published: January 2020

A soluble yellow CO dehydrogenase from CO-autotrophically grown cells of
Pseudomonas carboxydohydrogena
was purified 35-fold in seven steps to better than 95% homogeneity with a yield of 30%. The final specific activity was 180 μmol of acceptor reduced per min per mg of protein as determined by an assay based on the CO-dependent reduction of thionin. Methyl viologen, nicotinamide adenine dinucleotide (phosphate), flavin mononucleotide, and flavin adenine dinucleotide were not reduced by the enzyme, but methylene blue, thionin, and toluylene blue were reduced. The molecular weight of native enzyme was determined to be 4 × 10
5
. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate revealed at least three nonidentical subunits of molecular weights 14,000 (α), 28,000 (β), and 85,000 (γ). The ratio of densities of each subunit after electrophoresis was about 1:2:6 (α/β/γ), suggesting an α
3
β
3
γ
3
structure for the enzyme. The purified enzyme was free of formate dehydrogenase and nicotinamide adenine dinucleotide-specific hydrogenase activities, but contained particulate hydrogenase-like activity with thionin as electron acceptor. Known metalchelating agents tested had no effect on CO dehydrogenase activity. No divalent cations tested stimulated enzyme activity. The native enzyme does not contain Ni since cells assimilated little
63
Ni during growth, and the specific
63
Ni content of the enzyme declined during purification. The isoelectric point of the native enzyme was found to be 4.5 to 4.7. The

K
m

for CO was found to be 63 μM. The spectrum of the enzyme and its protein-free extract revealed that it contains bound flavin. The cofactor was flavin adenine dinucleotide based on enzyme digestion and thin-layer chromatography. One mole of native enzyme contains at least 3 mol of noncovalently bound flavin adenine dinucleotide.
Carbon monoxide metabolism of the methylotrophic acidogen Butyribacterium methylotrophicum
Authors: Lynd L , Kerby R , Zeikus J G
Published: January 2020

The Marburg strain of Butyribacterium methylotrophicum did not grow on CO alone but did consume CO during growth on a variety of substrates in the presence of a 100% CO gas phase. We selected a strain (the CO strain) that grew vigorously on CO alone. The ability of the CO strain to grow on CO was stable through multiple transfers in the absence of CO. CO dehydrogenase activity was lower in the CO strain grown on CO (13.3 micromol/min per mg of protein) than in the Marburg strain grown on methanol-acetate (47.2 mumol/min per mg of protein); thus, the levels of this enzyme did not explain the growth on CO. CO was dissimilated to acetate and CO2 with the following stoichiometry: 4 CO leads to 2.17 CO2 + 0.74 acetate. We observed a growth rate of 0.05 h-1, a final optical density at 660 nm of 0.8, and a cell yield of 3.0 g of cells per mol of CO during growth of the CO strain. Growing cultures of the CO strain displayed a Ks for CO of 28 to 56 microM. The apparent thermodynamic efficiency of cell synthesis from CO was 57%. Energetic and biochemical aspects of CO metabolism are described.
Induction of 3-Hydroxybenzoate 2-Hydroxylase in a
Pseudomonas testosteroni
Mutant
Authors: Daumy G. O. , McColl A. S.
Published: January 2020

Benzoate was established as the inducer of a unique 3-hydroxybenzoate 2-hydroxylase activity found in a
Pseudomonas testosteroni
mutant which is unable to grow on
m
-hydroxybenzoate as its sole source of carbon and energy.
Synthesis of acetyl coenzyme A from carbon monoxide, methyltetrahydrofolate, and coenzyme A by enzymes from Clostridium thermoaceticum
Authors: Hu S I , Drake H L , Wood H G
Published: January 2020

Two purified fractions from Clostridium thermoaceticum are shown to catalyze the following reaction: CO + CH3THF + CoA ATP leads to CH3COCoA + THF. The methyltetrahydrofolate (CH3THF) gives rise to the methyl group of the acetyl-coenzyme A (CoA) and the carbon monoxide (CO) and CoA to its carboxyl thio ester group. The role of ATP is unknown. One of the protein fractions (F2) is a methyltransferase, whereas the other fraction (F3) contains CO dehydrogenase and a methyl acceptor which is postulated to be a corrinoid enzyme. The methyltransferase catalyzes the transfer of the methyl group to the methyl acceptor, and the CO is converted to a formyl derivative by the CO dehydrogenase. By a mechanism that is as yet unknown, the formyl derivative in combination with CoA and the methyl of the methyl acceptor are converted to acetyl-CoA. It is also shown that fraction F3 catalyzes the reversible exchange of 14C from [1-14C]acetyl-CoA into 14CO and that ATP is required, but not the methyltransferase. It is proposed that these reactions are part of the mechanism which enables certain autotrophic bacteria to grow on CO. It is postulated that CH3THF is synthesized from CO and tetrahydrofolate which then, as described above, is converted to acetyl-CoA. The acetyl-CoA then serves as a precursor in other anabolic reactions. A similar autotropic pathway may occur in bacteria which grow on carbon dioxide and hydrogen.
Structural analysis of plasmid pSN2 in Staphylococcus aureus: no involvement in enterotoxin B production
Authors: Khan S A , Novick R P
Published: January 2020

Earlier studies have suggested the involvement of a small 1.3-kilobase plasmid, pSN2, in the production of enterotoxin D by certain Staphylococcus aureus strains. On the basis of extensive biochemical studies on pSN2, including the determination of its coding properties and its primary nucleotide sequence, we conclude that this plasmid is not in act involved in enterotoxin B production in S. aureus: although the toxin genes are apparently chromosomal, it is probable that they are part of a special genetic system such as a hitchhiking transposon.
Isolation and characterization of Pseudomonas aeruginosa R' plasmids constructed by interspecific mating
Authors: Morgan A F
Published: January 2020

Plasmid R68.45 was used to construct R' plasmids carrying a maximum of 4 to 5 map minutes of the Pseudomonas aeruginosa PAO chromosome by interspecific mating, using P. putida PPN as the recipient. These R' plasmids were used to determine the map location of the amiE locus and to identify tentatively a number of P. putida auxotrophic mutations. Some of these R' plasmids could not be maintained in recombination-deficient P. aeruginosa strains.
Extracellular exonuclease as a stage 0 biochemical marker in Bacillus subtilis sporulation
Authors: Burke W F , Slinker B S
Published: January 2020

Calcium-dependent exonuclease activity was produced by sporulating cells of Bacillus subtilis 168. Nuclease activity was released into the culture medium at approximately the same time as sporulation proteases, and production of these enzymes was tied to DNA replication. Results suggest that nuclease production is a function of the spo0H locus.
Regulation of purE transcription in a purE::lac fusion strain of Escherichia coli
Authors: Levine R A , Taylor M W
Published: January 2020

A purE::lac fusion strain was isolated by using a special Mu phage developed by M. Casadaban. In the presence of adenine (100 micrograms/ml), beta-galactosidase synthesis was repressed by greater than 90%. beta-Galactosidase activity could be detected 6 to 8 min after the removal of adenine and increased linearly for at least 20 min. purR- mutants were isolated and synthesized 1.7- to 1.8-fold-higher levels of beta-galactosidase compared with purR+ cells. Azaserine derepressed purE transcription approximately 1.7-fold by lowering purine nucleotide pools. Glutamine and pyrimidine supplementation or starvation had no effect on purE transcription. A comparison of the rate of de novo purine biosynthesis and purE transcription indicated that the in vivo rate of de novo purine biosynthesis was more sensitive to the inhibitory effects of adenine than was transcription at the purE locus.
Regulation of tryptophan genes in Rhizobium leguminosarum
Authors: Holmgren E , Crawford I P
Published: January 2020

Twelve tryptophan auxotrophs of Rhizobium leguminosarum were characterized biochemically. They were grown in complex and minimal media with several carbon sources, in both limiting and excess tryptophan. Missing enzyme activities allowed assignment of all mutant to the trpE, trpD, trpB, or trpA gene, confirming earlier results with the same mutants (Johnston et al., Mol. Gen. Genet. 165:323-330, 1978). In regulatory experiments, only the first enzyme of the pathway, anthranilate synthase, responded (about 15-fold) to tryptophan excess or limitation.
Partial nucleotide limitation induces phosphodiesterase I and 5'-nucleotidase in Bacillus subtilis
Authors: Dhariwal K R , Vasantha N , Freese E
Published: January 2020

Changes in the specific activity of enzymes involved in the degradation of RNA and nucleotides were measured in Bacillus subtilis under conditions of guanine deprivation, which initiates sporulation, and uracil deprivation, which does not initiate sporulation. Whereas the specific activities of most of the enzymes studied increased by less than a factor of 3, those of 5'-mononucleotide-producing phosphodiesterase and 5'-nucleotidase increased at least eightfold under both deprivation conditions.
Rhodopseudomonas sphaeroides membranes: alterations in phospholipid composition in aerobically and phototrophically grown cells
Authors: Onishi J C , Niederman R A
Published: January 2020

The effects of growth conditions on phospholipid composition in Rhodopseudomonas sphaeroides have been reexamined. The levels of phosphatidylethanolamine (27 to 28%), phosphatidylglycerol (23 to 24%), and phosphatidylcholine (11 to 18%) were very similar in cells grown aerobically or phototrophically at a high light intensity, consistent with findings for another member of Rhodospirillaceae. In addition, an unknown phospholipid species was detected which comprised 20 to 30% of the total phospholipid in these cells. In cells growing phototrophically at low-intensity illumination, the level of phosphatidylethanolamine increased by about 1.6-fold and that of the unknown phospholipid markedly decreased. Although the synthesis of photosynthetic pigments, light-harvesting protein, and intracytoplasmic photosynthetic membranes also increased markedly, the ratios of individual phospholipid species were essentially identical in photosynthetic membrane and cell wall fractions purified from these cells. Since a significant exchange of lipids apparently did not occur during the isolation of these fractions, it was suggested that the changes in cellular phospholipid accumulation were not due to a unique composition within the photosynthetic membrane. Instead, these phosphoglyceride changes were found to be related to overall phospholipid metabolism and could be accounted for principally by differences in biosynthetic rates. These results, together with studies in nutrient-restricted aerobic cells, suggested that the mechanism by which phospholipid levels are regulated may be related to radiant energy flux rather than cellular energy limitation.
TOL plasmid pWW0 in constructed halobenzoate-degrading Pseudomonas strains: enzyme regulation and DNA structure
Authors: Jeenes D J , Reineke W , Knackmuss H J , Williams P A
Published: January 2020

WR211 and WR216 are derivatives of halobenzoate-degrading Pseudomonas sp. strain B13 into which the 117-kilobase TOL degradative plasmid pWW0 has been transferred from Pseudomonas putida mt-2. WR211 has lost the ability to grow on the TOL-specific substrate m-xylene but retains the ability to grow on its metabolite, m-toluate. An analysis of the induction of enzymes was consistent with WR211 carrying a nonfunctional regulatory gene, xy1R, WR216 is a spontaneous derivative of WR211 which grows on one of the TOL substrates and yet expresses the nonspecific toluate oxidase, which enables it to grow on the novel substrate 4-chlorobenzoate. In addition to the xy1R lesion inherited from WR211, WR216 appears to carry a mutation in the structural gene for catechol 2,3-oxygenase, xy1E. The plasmids in both strains were analyzed by restriction endonuclease digestion. pWW0-1211 in WR211 has a large deletion (39 kilobases) compared with pWW0 and appears to be identical to a previously described plasmid (pWW0-8) which encodes none of the TOL degradative functions. pWW0-1216 in WR216 has undergone a major structural reorganization relative to its parent, pWW0-1211. This plasmid has a smaller deletion (19 kilobases), which is staggered relative to the deletion in pWW0-1211, and in addition it has two 3-kilobase insertions of unknown origin, one of which appears to cause the xylE mutation.
Effects of glnL and other regulatory loci on regulation of transcription of glnA-lacZ fusions in Klebsiella aerogenes
Authors: Goldie H , Magasanik B
Published: January 2020

Mutants of Klebsiella aerogenes containing genetic fusions of glnA to lacZ were isolated by using Mu dl (lac, bla) bacteriophage and a Mu Kmr helper phage with the host range of bacteriophage P1. Synthesis of beta-galactosidase in these strains is regulated in response to nitrogen metabolites and regulatory gln loci and is rendered constitutive by a mutation in the linked glnL gene. Complementation studies indicated that glnL is a separate locus from glnA and glnG and that insertions in glnA are partially polar on glnL expression. These results support the hypothesis that glnA, glnL, and glnG are organized in an operon with multiple promoters.
Ornithine metabolism in the genus Rochalimaea
Authors: Weiss E , Mamay H K , Dasch G A
Published: January 2020

Ornithine metabolism was studied in two strains of the trench fever rickettsia Rochalimaea quintana, Fuller and Guadalupe, and in the vole agent, a strain of Rochalimaea but not necessarily of Rochalimaea quintana. The metabolic activity of intact cells and cell-free extracts was measured by monitoring the evolution of 14CO2 from [1-14C]ornithine. Low levels of activity were obtained with all three strains, but requirements for the demonstration of this activity differed. With the cells of the Fuller and Guadalupe strains, the decarboxylation of ornithine was almost completely dependent on added pyruvate or succinate, presumably as sources of energy for transport. This enhancement was not prevented by the presence of chloramphenicol. The activity of the vole agent, on the other hand, required the complete medium. This activity was prevented by chloramphenicol added at the same time as the medium but not by chloramphenicol added after 1 h of incubation. In cell-free extracts, the demonstration of ornithine decarboxylase activity in the vole agent required prior induction with medium containing ornithine, whereas in the other two strains, the activity was constitutive. The activities of the extracts of the Fuller strain and the vole agent differed also in pH optimum, which was somewhat lower for the vole agent, and in the added pyridoxal phosphate requirement, which was greater for the Fuller strain. Comparable experiments with Rickettsia typhi and Rickettsia prowazekii failed to reveal evidence of ornithine metabolism.
Trailing flagella rotate faster than leading flagella in unipolar cells of Spirillum volutans
Authors: Swan M A
Published: January 2020

In unipolar cells of Spirillum volutans, the flagellar rotation frequency is halved, approximately, when the flagellar bundle reorientates to rotate about the cell body and reverse the swimming direction. The viscous drag resulting from a concomitant increase in flagellar wave amplitude is probably responsible for the reduced frequency of flagellar rotation.
Attachment of the chromosomal terminus of Bacillus subtilis to a fast-sedimenting particle
Authors: Sargent M G , Bennett M F
Published: January 2020

After gently lysed protoplasts of exponential phase cells of Bacillus subtilis were treated with restriction endonuclease BamHI, 99% of the DNA did not sediment with the plasma membrane. This DNA was fractionated on sucrose gradients into (i) a fast-sedimenting fraction highly enriched for genes from the origin and terminus (purA and ilvA), (ii) a 50 to 100S component also enriched for purA and ilvA, and (iii) the bulk of the DNA. The fast-sedimenting fraction was dissociated by Sarkosyl; this fraction contained a substantial amount of protein and is probably a membrane subparticle. The S value of the 50 to 100S component was not greatly affected by Sarkosyl treatment, but these particles were unable to penetrate an agarose gel during electrophoresis and were retained by nitrocellulose filters. The terminus DNA in the fast-sedimenting fraction and the 50 to 100S component contained a large restriction fragment (1.5 x 10(7) to 2.0 x 10(7) daltons) encoding ilvA, thyB, and ilvD. The bulk of the SP beta prophage and metB, which lie to the right and left, respectively, of the ilvA-ilvD cluster, were not part of the complex. citK, which lies to the right of SP beta, appeared to be present in the fast-sedimenting complexes. The neighboring genes kauA and gltA were not part of the fast-sedimenting complexes. The presence of terminus DNA in the fast-sedimenting components was also demonstrated by a radiochemical method.
Aryl-
l
-Aminoacylamidase Activities in Extracts of
Streptococcus durans
Authors: Machuga Edward J.
Published: January 2020

Two distinct enzymes with aryl-
l
-aminoacylamidase activity were found in cellular extracts of
Streptococcus durans
. One of these enzymes was strictly an arylamidase lacking any observable N-terminal exopeptidase activity. The other enzyme functioned as an aminopeptidase capable of catalyzing the hydrolysis of a variety of
l
-peptide and arylamide substrates. The arylamidase (molecular weight, 80,000) purified 425-fold to homogeneity preferred arylamides containing large hydrophobic side chains, whereas the partially purified aminopeptidase (molecular weight, 300,000) preferred substrates with small nonpolar or basic side chains. Neither enzyme contained any endopeptidase or carboxypeptidase activity. The purified arylamidase was unaffected by metal chelators, but Mn
2+
and Mg
2+
did act as nonessential activators exclusively affecting the maximal velocity. The arylamidase-catalyzed hydrolysis of
l
-leucyl-
p
-nitroanilide exhibited a bell-shaped pH dependence for log
V
max
/

K
m

(pK
1
of 7.2; pK
2
of 8.5), whereas the log
V
max
-versus-pH profile showed only an acid limb (pK of 6.8). The ionizable group responsible for the basic limb of the log
V
max
/

K
m

-versus-pH profile corresponded to the α-amino group of the substrate
l
-leucyl-
p
-nitroanilide (pK
a
= 8.5). Diazoacetyl-
dl
-norleucine methyl ester (2 mM) in the presence of 100 mM Cu
2+
caused a rapid inactivation of the enzyme (
t
½
of 24 min). Neither parachloromercuribenzoate (0.5 mM) nor
N
-ethylmaleimide (50 mM) had any effect on the arylamidase activity. Reversible noncompetitive inhibition was observed for iodoacetate (

K
i

of 30 mM),
N
-acetylimidazole (

K
i

of 4.0 mM), and ethyl acetimidate (

K
i

of 45 mM), although time-dependent irreversible inactivation was not observed with these reagents.
Effect of Intracytoplasmic Membrane Development on Oxidation of Sorbitol and Other Polyols by
Gluconobacter oxydans
Authors: White S. A. , Claus G. W.
Published: January 2020

By using membrane-bound dehydrogenases,
Gluconobacter oxydans
characteristically accomplishes single-step oxidation of many polyols and quantitative release of the oxidation product into the medium. These cells typically differentiate by forming intracytoplasmic membranes (ICM) after exponential growth on glycerol. Earlier experiments demonstrated that glycerol-grown cells containing ICM oxidized glycerol more rapidly than cells which were harvested during exponential growth and lacked ICM (Claus et al., J. Bacteriol.
123:
1169-1183). This report demonstrates that ICM are also formed after growth on sorbitol. Sorbitol-grown, ICM-containing maximum stationary-phase (MSP) cells showed from 50 to 300% greater oxidation (respiration) rates on mannitol, glycerol, glucose,
meso
-erythritol, and
meso
-inositol than did exponential-phase (EXP) cells which lacked ICM. Both EXP and MSP cells exhibited maximum sorbitol oxidation at pH 5.0, 38°C, and 5% (wt/vol) sorbitol. When assayed under these optimum conditions, ICM-containing MSP cells demonstrated a 72% increase in respiration on sorbitol compared with that of EXP cells lacking ICM (oxygen quotients of 3,100 and 1,800, respectively). Gas chromatographic studies showed that sorbose was the only detectable product released from cells during oxygen quotient analysis. The specific activity of particulate-bound sorbitol dehydrogenase from ICM-containing MSP cells was twice that obtained from particulate fractions prepared from EXP cells lacking ICM. These results show that neither ICM formation after exponential growth nor increased respiration of other polyols is dependent upon the polyol used to grow cells. Our results suggest that increased respiratory activity of MSP cells is caused both by ICM formation and by increased synthesis (or activity) of the polyol dehydrogenases found in these membranes.
Thiamine transport in Saccharomyces cerevisiae protoplasts
Authors: Nishimura H , Sempuku K , Iwashima A
Published: January 2020

Thiamine was found to be accumulated in protoplasts of Saccharomyces cerevisiae in the same manner as in intact cells, suggesting that a soluble thiamine-binding protein in periplasm may not be an essential component of the thiamine transport system of S. cerevisiae. It was also found that thiamine pyrophosphate cannot be taken up by yeast protoplasts.
Cadmium and manganese transport in Staphylococcus aureus membrane vesicles
Authors: Perry R D , Silver S
Published: January 2020

The presence of plasmid gene cadB did not affect Cd2+ accumulation, whereas plasmid gene cadA reduced Cd2+ accumulation by whole cells but not by membrane vesicles. Membrane vesicle studies indicated that Cd2+ uptake occurred via the Mn2+ transport system which was energized by the membrane electrical potential. Mn2+ and Cd2+ were competitive inhibitors of each other's transport, with Km's of 0.95 microM Mn2+ and 0.2 microM Cd2+. The kinetic parameters were nearly identical with vesicles prepared from sensitive and resistant cells, indicating that the cadA-encoded Cd2+ efflux system was inoperative in membrane vesicle preparations. Experiments with energy-inhibited cells indicated that the cadB gene product may bind Cd2+.
Induction of acyl coenzyme A synthetase and hydroxyacyl coenzyme A dehydrogenase during fatty acid degradation in Neurospora crassa
Authors: Hii V , Courtright J B
Published: January 2020

Neurospora crassa is able to use long-chain fatty acids as the sole carbon and energy source. After growth on oleate there was nearly a 10-fold induction of the acyl coenzyme A (CoA) synthetase and a fivefold increase in the activity of the 3-hydroxyacyl-CoA dehydrogenase. There was a slight induction of the enoyl-CoA hydratase and 3-ketoacyl-CoA thiolase, but no apparent induction of the flavin-linked acyl-CoA dehydrogenase. These noncoordinate changes in the fatty acid degradation enzymes suggest that they are not organized into a multienzyme complex as is found in bacteria.
Fructose 6-phosphate phosphorylation in Bacteroides species
Authors: Roberton A M , Glucina P G
Published: January 2020

6-Phosphofructokinase (6-PFK) activities have been measured in cell extracts from a number of Bacteroides species. Two main types of 6-PFK were found: an ATP-linked 6-PFK and a PPi-linked 6-PFK. In most strains both of these activities were found, although in two strains only ATP-linked 6-PFK was present. The PPi-linked 6-PFK activity was always higher, when both activities were present, and showed Michaelis-Menten kinetics with respect to fructose 6-phosphate. In contrast, the ATP-linked 6-PFK activity usually gave sigmoid kinetics with respect to fructose 6-phosphate, although several strains were found to have activities with Michaelis-Menten kinetics. Conditions for measuring maximum activities of ATP-linked 6-PFK were not identical for different strains, and the activities were rather unstable in extracts. The possible consequences of the observation that most Bacteroides strains possess both an ATP-linked and a PPi-linked 6-PFK are discussed.
The BtuB Group Col Plasmids and Homology Between the Colicins They Encode
Authors: Mock Michèle , Pugsley Anthony P.
Published: January 2020

Colicins A, E1, E2, E3, E4, E5, E6, and E7 exhibited reduced activity against BtuB mutants of
Escherichia coli
K-12 and also against wild-type cells in the presence of vitamin B12. Plasmids encoding representatives of these colicins were specifically immune to high levels of the homologous colicin. Col
+
cells grown in media containing mitomycin C accumulated large amounts of colicin polypeptide. ColE2
+
, ColE3
+
, ColE4
+
, ColE5
+
, and ColE6
+
cultures also synthesized large amounts of second, lower-molecular-weight protein under these conditions. Colicins E2 through E7, but not A or E1, reacted with antiserum raised against purified colicin E3. Colicins E2 and E7 induced synthesis of β-galactosidase encoded by
lacZ
under the control of the colicin Ib gene promotor on a derivative of Col plasmid ColIb.P9. This promotor is usually active only when the cells are treated with agents which damage DNA or block replication. Plasmids encoding various mutant forms of colicin E3 (M. Mock and M. Schwartz, J. Bacteriol.
142:
384-390, 1980) recombined with ColE2, ColE4, ColE5, or ColE6 plasmids at a frequency of 10
−4
per cell to produce a colicin active against ColE2
+
, E4
+
, E5
+
, or E6
+
cells. ColE5 and ColE6 plasmids recombined with ColE3 plasmids bearing mutations affecting colicin E3 receptor recognition, envelope penetration, and catalytic activities. ColE2 and ColE4 plasmids recombined only with ColE3 plasmids bearing mutations affecting receptor recognition and envelope penetration. Recombinants between mutant ColE3 plasmids and ColA, ColE1, or ColE7 plasmids were not detected. We propose the designation BtuB group for the colicins described here, and we divide the group into two classes comprising colicins A and E1, which act on the cytoplasmic membrane, and the related colicins E2 through E7, which have known or putative nuclease activities.
Origin and direction of mini-R1 plasmid DNA replication in cell extracts of Escherichia coli
Authors: Diaz R , Staudenbauer W L
Published: January 2020

Replication of the mini-R1 plasmids pKN177 and pKN182 can be carried out efficiently in cell extracts of Escherichia coli and depends on both transcription and translation. Heteroduplex and restriction analyses indicate that both plasmids are derived from the R1 copy mutant pKN104 by Is1-mediated recombination events without involving structural alterations in the replication region. To ascertain whether the in vitro replication of these miniplasmids corresponds to R1 replication in vivo, the origin and direction of replication were analyzed by electron microscopy of replicative intermediates. It was found that replication starts at a unique origin located within the RepA region at R1 coordinate at 82.4 kilobases and proceeds unidirectionally toward the IS/b sequence. The specification of the origin and the direction of in vitro replication are therefore in full agreement with the pattern observed previously for the in vivo replication of the closely related plasmids R100 and R6-5. This agreement provides additional evidence that R1 DNA synthesis in vitro employs the same replication mechanism as it does in vivo.
Chromosomal location of att P7, the recA-independent p7 integration site used in the suppression of Escherichia coli dnaA mutations
Authors: Chesney R H , Adler E
Published: January 2020

Conjugational and transductional analyses were used to determine the chromosomal location of attP7, the recA-independent integration site used by bacteriophage P7 to suppress host dnaA mutations. The site of integration was found to be between tolC and dnaG. An increase in transduction frequencies was observed for markers surrounding attP7 when P7 was integrated. Under these conditions, all pairs of markers in this region, including those separated by attP7, were cotransduced at frequencies higher than normal, indicating the possible production of P7 specialized transducing particles.
Nitrogen control in Pseudomonas aeruginosa: mutants affected in the synthesis of glutamine synthetase, urease, and NADP-dependent glutamate dehydrogenase
Authors: Janssen D B , Habets W J , Marugg J T , Van Der Drift C
Published: January 2020

Mutants were isolated from Pseudomonas aeruginosa that were impaired in the utilization of a number of nitrogen sources. In contrast to the wild-type strain, these mutants appeared to be unable to derepress the formation of glutamine synthetase and urease under nitrogen-limited growth conditions, whereas NADP-dependent glutamate dehydrogenase became derepressed. This GlnR- phenotype appeared to be caused by a mutation located in the early region of the P. aeruginosa PAO chromosomal map, close to hisIV59. Partial suppression of the GlnR- phenotype due to a mutation located close to hisII4 was observed. These revertants were different from both the wild-type strain and the GlnR- mutant with respect to the regulation of the synthesis of glutamine synthetase, urease, and NADP-dependent glutamate dehydrogenase (GlnRc phenotype). Also the regulation of glutamine synthetase activity by adenylylation/deadenylylation was altered in the revertants. The results suggest the presence of a regulatory gene that plays a role in the regulation of enzyme formation in response to the availability of ammonia.
Quantitative analysis of the heat shock response of Saccharomyces cerevisiae
Authors: Miller M J , Xuong N H , Geiduschek E P
Published: January 2020

Transient protein synthesis in Saccharomyces cerevisiae, after shift from 21-23 degrees C to 37 degrees C, was quantitatively analyzed. Pulse-labeled proteins were separated by two-dimensional gel electrophoresis, and autoradiograms of the gels were analyzed by a recently described method involving a computer-coupled film scanning system. In this way, the rate of incorporation of L-[35S]methionine into approximately 500 proteins was followed. The synthesis of more than 80 of these proteins was transiently induced at 37 degrees C, with about 20 being classified as major heat shock proteins (defined as those whose rate of labeling was increased at least eightfold at some time during the response). The synthesis of more than 300 of the proteins was transiently repressed at 37 degrees C, and several general temporal patterns of repression could be distinguished. The influence of temperature-sensitive mutations affecting RNA synthesis and transport on the heat shock response was also examined. A protein whose induction in response to heat shock has a post-transcriptional component could be identified. As previously pointed out, the heat shock repression of certain proteins is so rapid that it also must involve post-transcriptional effects.
Metabolism of dimethylphthalate by Micrococcus sp. strain 12B
Authors: Eaton R W , Ribbons D W
Published: January 2020

During growth of Micrococcus sp. strain 12B with dimethylphthalate, 4-carboxy-2-hydroxymuconate lactone (CHML, X) and 3,4-dihydroxyphthalate-2-methyl ester (XI) were isolated from culture filtrates. CHML is the lactone of intermediate 4-carboxy-2-hydroxymuconate (IX). Accumulation of XI which is not a substrate for 3,4-dihydroxyphthalate-2-decarboxylase in strain 12B afforded an easy access to the preparation of 3,4-dihydroxyphthalate.
Phosphoenolpyruvate-dependent phosphotransferase system enzyme III and plasmid-encoded sucrose transport in Escherichia coli K-12
Authors: Lengeler J W , Mayer R J , Schmid K
Published: January 2020

The phosphoenolpyruvate-dependent carbohydrate:phosphotransferase system enzyme IISCR, specific for and regulated by sucrose, was analyzed in derivatives of Escherichia coli K-12 carrying the sucrose plasmid pUR404. Enzyme IIScr, coded for by gene scrA of the plasmid, depended for its transport and phosphorylation activity directly on the phosphotransferase system enzyme IIIGlc, Scr, coded for by the chromosomal gene crr.
RNA polymerase of Myxococcus xanthus: purification and selective transcription in vitro with bacteriophage templates
Authors: Rudd K E , Zusman D R
Published: January 2020

DNA-dependent RNA polymerase from vegetative cells of the gram-negative, fruiting bacterium Myxococcus xanthus was purified more than 300-fold by a modified Burgess procedure (Lowe et al., Biochemistry 18:1344-1352, 1979), using Polymin P precipitation, 40 to 65% saturated ammonium sulfate fractional precipitation, double-stranded DNA cellulose chromatography, A5m gel filtration chromatography, and single-stranded DNA agarose chromatography. The last step separated the RNA polymerase into a core fraction and an enriched holoenzyme fraction. The core enzyme showed a subunit structure similar to that of the Escherichia coli polymerase, as follows: beta' and beta (145,000 and 140,000 daltons, respectively) and alpha (38,000 daltons). A comparison of the core enzyme and the holoenzyme implicated two polypeptides as possible sigma subunits. These polypeptides were closely related, as indicated by peptide analysis. M. xanthus RNA polymerase was capable of transcribing DNAs from E. coli phages T7, T4, and lambda, Bacillus subtilis phage phi 29, and M. xanthus phages Mx1, Mx4, and Mx8. Transcription of T7 and phi 29 DNAs was stimulated by KCl, whereas transcription of Mx1, Mx4, and Mx8 DNAs was inhibited by KCl. Magnesium ion dependence, rifampin and heparin sensitivities, and spermidine stimulation of M. xanthus RNA polymerase activity were similar to those found with E. coli RNA polymerase. The pH optimum of M. xanthus RNA polymerase activity was more basic than that of E. coli polymerase. M. xanthus RNA polymerase was capable of selective transcription in vitro when DNAs from phages T7 delta 111, phi 29, and Mx1 were used. The molecular weights of the resulting phage RNA transcripts made by M. xanthus RNA polymerase (as determined by agarose-acrylamide slab gel electrophoresis) were the same as the molecular weights of the transcripts synthesized by E. coli RNA polymerase. No discrete transcripts were detected as the in vitro RNA products of M. xanthus phage Mx4 and Mx8 DNA transcription. Southern transcript synthesized by M. xanthus RNA polymerase. Three transcripts (transcripts A, B, and C; molecular weights, 2.55 X 10(6), 1.95 X 10(6), and 1.56 X 10(6), respectively) were identified as in vitro RNA products of M. xanthus phage Mx1 DNA transcription when either E. coli or M. xanthus RNA polymerase was used. A Southern blot hybridization analysis indicated that the E. coli RNA polymerase and the M. xanthus RNA polymerase transcribe common SalI restriction fragments of Mx1 DNA.
Comparison of DNA sequences required for the function of citrate utilization among different citrate utilization plasmids
Authors: Shinagawa M , Makino S , Hirato T , Ishiguro N , Sato G
Published: January 2020

The relatedness of DNA sequences encoding citrate utilization was examined by hybridization with a cloned DNA fragment from a citrate utilization (Cit) plasmid, pOH30221, and DNA of other Cit plasmids. This revealed that there are at least two groups of Cit plasmids: the Inc W Cit plasmids, which show homology with the probe, and the Inc H1 plasmids, which do not.
Transposition of Tn7 Occurs at a Single Site on the
Caulobacter crescentus
Chromosome
Authors: Ely Bert
Published: January 2020

Insertion of Tn7 in nine independently derived isolates occurred in a single small region of the
Caulobacter crescentus
chromosome, suggesting that Tn7 insertion occurs at or near a single site. The order of markers in this region of the
Caulobacter
chromosome was shown to be
flaA-argG
-Tn7.
Chromosomal genes for ColV plasmid-determined iron(III)-aerobactin transport in Escherichia coli
Authors: Braun V , Burkhardt R , Schneider R , Zimmermann L
Published: January 2020

Four chromosomal genes, tonA (fhuA), fhuB, tonB, and exbB, were required for the transport of iron(III)-aerobactin specified by the plasmids ColV-K311, ColV-K229, ColV-K328, and ColV-K30. These genes also determine the transport system in Escherichia coli for the iron ionophore ferrichrome. Aerobactin and ferrichrome are both iron ligands of the hydroxamate type, but they are of different structure. The ColV plasmids determine an outer membrane protein that serves as a receptor for cloacin. Cloacin-resistant mutants were devoid of iron(III)-aerobactin transport but were unimpaired in ferrichrome transport. We conclude that for iron(III)-aerobactin transport two outer membrane proteins, the TonA and the cloacin receptor protein, have to interact functionally or structurally or both.