bluuebunny/crossref_metadata_2025 huggingface dataset:
Inhibition and repression of homocitrate synthase by lysine in Penicillium chrysogenum
Authors: Luengo J M , Revilla G , López M J , Villanueva J R , Martín J F
Published: January 2020

Homocitrate synthase in the first enzyme of the lysine biosynthetic pathway. It is feedback regulated by L-lysine. Lysine decreases the biosynthesis of penicillin (determined by the incorporation of [14C]valine into penicillin) by inhibiting and repressing homocitrate synthase, thereby depriving the cell of alpha-aminoadipic acid, a precursor of penicillin. Lysine feedback inhibited in vivo the biosynthesis and excretion of homocitrate by a lysine auxotroph, L2, blocked in the lysine pathway after homocitrate. Neither penicillin nor 6-aminopenicillanic acid exerted any effect at the homocitrate synthase level. The molecular mechanism of lysine feedback regulation in Penicillium chrysogenum involved both inhibition of homocitrate synthase activity and repression of its synthesis. In vitro studies indicated that L-lysine feedback inhibits and represses homocitrate synthase both in low- and high-penicillin-producing strains. Inhibition of homocitrate synthase activity by lysine was observed in cells in which protein synthesis was arrested with cycloheximide. Maximum homocitrate synthase activity in cultures of P. chrysogenum AS-P-78 was found at 48 h, coinciding with the phase of high rate of penicillin biosynthesis.
Formation of a lambda (Tn10) tyrR+ specialized transducing bacteriophage from Escherichia coli K-12
Authors: Cobbett C S , Pittard J
Published: January 2020

The transposon Tn10, coding for resistance to tetracycline, was inserted close to the tyrR+ gene at min 28 on the Escherichia coli chromosome. The homology between this transposon and a lambda (Tn10) phage was employed to direct integration of lambda close to tyrR+ with subsequent isolation of a lambda (Tn10) tyrR+ transducing phage. Results of restriction endonuclease analysis of the transducing phage are presented.
Genetic transformation with cell wall-associated deoxyribonucleic acid in Bacillus subtilis
Authors: Doyle R J , Streips U N , Imada S , Fan V S , Brown W C
Published: January 2020

Cell walls from bacillus subtilis 168 were prepared by conventional methods and found to contain deoxyribonucleic acid (DNA). In transformation assays, after autolysis, it was found that two major regions of the chromosome were selectively enriched in the wall preparations. One region clustered around the replication origin and is represented by the markers purA16, ts8132, thiC5, sacA321, and hisA1. The other region included the replication terminus with representative loci metB10, citK5, gltA292, and pyrA1. All other (internal) loci which were examined showed no statistical enrichment. The two areas of enrichment were similar to but more extensive than those reported for membrane-DNA complexes. The wall preparations also contained protein and lipid, indicating a possible membrane involvement. Analyses of the cell walls revealed that the fatty acid composition of the membrane component was not typical of the for B. subtilis protoplast membranes or for lipoteichoic acids. In addition, radioiodination of cell wall autolysates, followed by gel electrophoresis and autoradiography, demonstrated the presence of proteins not readily detectable in bulk protoplast membranes or on the surfaces of intact cells. These data suggest that a unique component of the membrane and regions of the B. subtilis genome involved in DNA replication events are tightly associated with cell walls. The binding of DNA-membrane complexes to the "rigid" cell wall and the replication of the wall could be a mechanism by which the segregation of growing chromosomes occurs.
Regulation and glutamic acid decarboxylase during Neurospora crassa conidial germination
Authors: Christensen R L , Schmit J C
Published: January 2020

Glutamic acid decarboxylase (GAD) from Neurospora crassa was assayed in dormant and germinating conidia that had been permeabilized by toluene and methanol. N. crassa conidia contained 10 times the GAD activity found in vegetativemycelia. During conidial germination, GAD activity rapidly decreased to low levels before germ tubes appeared. GAD activity in germinating conidia closely followed the decreasing rate of glutamic acid metabolism. Inhibiting protein synthesis partially blocked the decrease in GAD activity, but eliminating exogenous carbon sources did not alter the initial rate of decrease in this enzyme. However, when conidia were incubated for more than 3 h in distilled water, GAD activity began to increase and eventually reached levels comparable to those in dormant conidia. Either GAD was reversibly inactivated or this enzyme could be synthesized from endogenous storage compounds when conidia were incubated in distilled water. These results are consistent with the hypothesis that GAD is a developmentally regulated enzyme that is responsible for catalyzing the first step in the metabolism of the large pool of free glutamic acid during conidial germination.
Genetic regulation of variable Vi antigen expression in a strain of Citrobacter freundii
Authors: Snellings N J , Johnson E M , Kopecko D J , Collins H H , Baron L S
Published: January 2020

Certain strains of the genus Citrobacter exhibit a variable expression of the Vi surface antigen that appears to involve a special mechanism for regulation of gene expression. Two nonlinked chromosomal loci, viaA and viaB, are known to determine nonvariable Vi antigen expression in strains of Salmonella. To confirm the presence of analogous loci in Citrobacter and to ascertain whether either of them is involved in variable Vi antigen expression in this organism, donor strains were constructed from Citrobacter freundii WR7004 and used to transfer their Vi antigen-determining genes to ViaA- and ViaB- Salmonella typhi recipient strains. Vi antigen expression in C. freundii was found to be controlled by loci analogous to the Salmonella via genes. S. typhi recipients of the C. freundii viaA+ genes were restored to the full, continuous expression of the Vi antigen normally seen in S. typhi. Thus, the C. freundii viaA genes appeared to play no role in the variable expression of the Vi antigen. In contrast, S. typhi recipients of the C. freundii viaB+ genes exhibited the rapid, reversible alternation between full Vi antigen expression and markedly reduced Vi antigen expression that was seen to occur in the C. freundii parent. The C. freundii viaB locus was thus identified as the one whose genes are regulated so as to produce variable Vi antigen expression. Genes determining another C. freundii surface antigen, the synthesis of which is not affected by the mechanism regulating Vi expression, were coinherited with the C. freundii viaB+ genes. An invertible, insertion sequence element located within the C. freundii viaB locus is proposed to account for the regulation of variable Vi antigen expression.
Plasmid transfer within and between serologically distinct strains of Rhizobium japonicum, using antibiotic resistance mutants and auxotrophs
Authors: Pilacinski W P , Schmidt E L
Published: January 2020

Methionine-requiring and pantothenic acid-requiring auxotrophs of Rhizobium japanicum USDA 31, as well as highly antibiotic-resistant mutants of R. japonicum strains USDA 31, USDA 110, USDA 138, and Webster 48, were isolated. These mutants were used to transfer the P-1 group plasmids R68.45 and RP4 within and between strains USDA 31, USDA 110, and Webster 48. Attempts to demonstrate transfer of either plasmid to strain USDA 138 were unsuccessful.
Specialized transducing bacteriophage lambda carrying the structural gene for a major outer membrane matrix protein of Escherichia coli K-12
Authors: Mutoh N , Nagasawa T , Mizushima S
Published: January 2020

A specialized transducing phage lambda carrying the structural gene for the OmpF protein, an outer membrane matrix protein, was isolated. The phage carries the 20.5--21-min region of the Escherichia coli K-12 chromosome and carries asnS, ompF, and aspC genes.
A Salmonella typhimurium mutant dependent upon carbamyl aspartate for resistance to 5-fluorouracil is specifically affected in ubiquinone biosynthesis
Authors: Zak V L , Kelln R A
Published: January 2020

The isolation and properties of a mutant dependent upon exogenous carbamyl aspartate for resistance to 5-fluorouracil are described. The mutant was deficient in the synthesis of ubiquinone and accumulated a quinone provisionally identified as the ubiquinone precursor 2-octaprenyl-3-methyl-6-methoxy-1,4-benzoquinone. The mutation resulted in an alteration in the regulation of synthesis of enzymes involved in de novo pyrimidine biosynthesis but did not establish a functional block in dihydroorotate dehydrogenase activity in vivo. Conditional resistance to 5-fluorouracil apparently occurred through an inhibition of the conversion of the analog to the nucleotide level.
Plasmid transformation in Haemophilus influenzae
Authors: Albritton W L , Bendler J W , Setlow J K
Published: January 2020

Purified 34-megadalton-plasmid deoxyribonucleic acid from antibiotic-resistant strains of Haemophilus influenzae transforms competent strains of H. influenzae more efficiently if the recipient strains contain certain other 30-megadalton plasmids.
Host requirements for growth of lambda-P22 hybrid in Escherichia coli
Authors: Taylor K D , Shizuya H
Published: January 2020

The requirements for growth of bacteriophage lambda containing the deoxyribonucleic acid replication region from Salmonella phage P22 were determined in a burst size experiment. The products of genes dnaE, dnaJ, dnaK, dnaY, dnaZ, and seg were required, but not the products of genes dnaA, dnaB, dnaC, and dnaX. This lambda-P22 hybrid phage was also dependent on polA for growth at 32 degrees C.
Lipid metabolism during encystment of Azotobacter vinelandii
Authors: Reusch R N , Sadoff H L
Published: January 2020

The formation of cysts by Azotobacter vinelandii involves the synthesis of lipids as major metabolic products. Cells which encyst at low levels in aging glucose cultures undergo the same pattern of lipid synthesis as cells which undergo reasonably synchronous encystment in beta-hydroxybutyrate or n-butanol. The accumulation of poly-beta-hydroxybutyrate (PHB) precedes the synthesis of 5-n-heneicosylresorcinol and 5-n-tricosylresorcinol (AR1), which is then followed in about 6 h by the synthesis of the 5-n-alkylresorcinol galactosides (AR2). In the mature cyst, PHB, AR1, and AR2 account for 8, 5.6, and 4.5%, respectively, of the dry weight. Phospholipid formation levels off 4 h postinduction, which coincides with the final cell division, but fatty acids synthesis continues at a very low level throughout encystment, suggesting some turnover of fatty acid. Distribution studies show that AR1 and AR2 are found in roughly equal amounts in the exine and central body of the cysts, with only trace amounts recovered from the intine. Studies of cysts labeled during encystment with [14C]beta-hydroxybutyrate or during vegetative growth with [14C]glucose suggest that the exine structure is synthesized during encystment, but that the intine is composed largely of vegetative cell components.
Molecular cloning of TOL genes xylB and xylE in Escherichia coli
Authors: Inouye S , Nakazawa A , Nakazawa T
Published: January 2020

The xylB and xylE genes in the TOL plasmid of Pseudomonas putida mt-2, which code for benzyl alcohol dehydrogenase and catechol 2,3-oxygenase, respectively, were cloned onto plasmid pBR322 in Escherichia coli for detailed mapping. The xylB gene was mapped in a 2.9-kilobase region within the BamHI BC fragment of pTN2, an in vivo RP4-TOL recombinant, whereas the xylE gene was mapped in a 1.8-kilobase region within the BamHI BD fragment. The directions of transcription of these genes were deduced from the expression of the cloned genes which had been ligated in orientations opposite pBR322 at its BamHI site within the tetracycline resistance gene. The xylB and xylE genes are inducible by a specific inducer of the TOL pathway genes in the RP4-TOL recombinant, whereas they are not inducible in the pBR322-TOL hybrids. The regulatory regions involved in expression of the xylB and xylE genes do not appear to be located in the vicinity of the structural genes. Catechol 2,3-oxygenase formed in E. coli carrying an xylE-containing plasmid is similar, or identical, to that formed in P. putida carrying the TOL plasmid.
In vivo intermembrane transfer of phospholipids in the photosynthetic bacterium Rhodopseudomonas sphaeroides
Authors: Cain B D , Deal C D , Fraley R T , Kaplan S
Published: January 2020

The kinetics of accumulation of phospholipids into the intracytoplasmic membrane of Rhodopseudomonas sphaeroides have been examined. We have previously demonstrated that accumulation of phospholipids in the intracytoplasmic membrane is discontinuous with respect to the cell cycle. In this study we demonstrated a sevenfold increase in the rate of phospholipid incorporation into the intracytoplasmic membrane concurrent with the onset of cell division. Pulse-chase labeling studies revealed that the increase in the rate of phospholipid accumulation into the intracytoplasmic membrane results from the transfer of phospholipid from a site other than the intracytoplasmic membrane, and that the transfer of phospholipid, rather than synthesis of phospholipid, is most likely subject to cell cycle-specific regulation. The rates of synthesis of the individual phospholipid species (phosphatidylethanolamine, phosphatidyglycerol, and an unknown phospholipid) remained constant with respect to one another throughout the cell cycle. Similarly, each of these phospholipid species appeared to be transferred simultaneously to the intracytoplasmic membrane. We also present preliminary kinetic evidence which suggested that phosphatidylethanolamine may be converted to phosphatidycholine within the intracytoplasmic membrane.
Topographical distribution of penicillin-binding proteins in the Escherichia coli membrane
Authors: Buchanan C E
Published: January 2020

The penicillin-binding proteins (PBPs) found in the membranes of Escherichia coli X925 minicells (primarily cell ends or septa) were compared with those found in rod-shaped cells (primarily sidewalls) in an effort to determine whether certain PBPs are unevenly distributed over the bacterial cell membrane. The seven major PBPs of E. coli were all present in minicell membranes. PBP 1B was altered in minicells, however, appearing as two bands on sodium dodecyl sulfate-polyacrylamide gels rather than the usual three. PBP 2, which is needed for longitudinal growth of the cell but not for septum formation, was significantly reduced in minicell membranes. This observation is consistent with the fact that minicells contain very little sidewall material and raises the possibility that the specialized function of PBP 2 may be determined or regulated by its uneven topographical distribution in the membrane. None of the PBPs appeared to be selectively enriched in minicell membranes.
Timing of ribosome synthesis during ascosporogenesis of yeast cells: evidence for early function of haploid daughter genomes
Authors: Emanuel J R , Magee P T
Published: January 2020

During meiosis and sporulation in Saccharomyces cerevisiae, the recessive genetic marker for cycloheximide resistance, believed to be due to an altered ribosomal protein (C. S. McLaughlin, p. 815-827, in M. Nomura et al., ed., Ribosomes, Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y.), is expressed as early as meiosis II. Ribosomal ribonucleic acid synthesis peaks near the time that cycloheximide resistance begins to appear. Less than 25% of the 17S and 25S ribonucleic acid of the vegetative cells persists in spores, but pulse-labeling studies indicate that greater than 90% of the stable ribonucleic acid made after 6 h survives in spores. These results indicate that the haploid daughter genomes begin to function near the time of meiosis II.
Phospholipid biosynthesis is required for stalk elongation in Caulobacter crescentus
Authors: Mansour J D , Henry S , Shapiro L
Published: January 2020

Membrane phospholipid synthesis was inhibited in Caulobacter crescentus by growth of a glycerol auxotroph in the absence of glycerol or by treatment with the antibiotic cerulenin. It was observed that the final step in the swarmer cell-to-stalked cell transition, stalk elongation, was inhibited under these conditions. Since an early effect of inhibiting phospholipid synthesis in C. crescentus is the termination of deoxyribonucleic acid (DNA) replication (I. Contreras, R. Bender, A. Weissborn, K. Amemiya J. D. Mansour, S. Henry, and L. Shapiro, J. Mol. Biol. 138:401-410, 1980), we questioned whether the inhibition of stalk formation was due directly to the inhibition phospholipid synthesis or secondarily to the inhibition of DNA synthesis. Under conditions which inhibited DNA synthesis but permitted phospholipid synthesis, i.e., growth of a temperature-sensitive DNA elongation mutant at the restrictive temperature or treatment with hydroxy-urea, stalk elongation occurred normally. Therefore phospholipid synthesis is required for stalk elongation in C. crescentus.
Two succinic semialdehyde dehydrogenases are induced when Escherichia coli K-12 Is grown on gamma-aminobutyrate
Authors: Donnelly M I , Cooper R A
Published: January 2020

When Escherichia coli K-12 was grown on gamma-aminobutyrate, a second succinic semialdehyde dehydrogenase, dependent upon oxidized nicotinamide adenine dinucleotide or oxidized nicotinamide adenine dinucleotide phosphate and distinct from that induced by gamma-aminobutyrate, was gratuitously induced by succinic semialdehyde.
Properties of the lactose transport system in Klebsiella sp. strain CT-1
Authors: Imai K , Hall B G
Published: January 2020

Highly purified [D-glucose-1-14C]lactose has been used to study the transport of lactose by Klebsiella sp. strain CT-1. Strain CT-1 transports lactose by a lactose-inducible system that exhibited an apparent Km of 6 mM lactose and an apparent Vmax of 140 nmol/min per mg of cell protein. Lactose uptake was inhibited competitively by o-nitrophenyl-beta-D-galactoside with a Ki value of 8 mM, but was not inhibited by thio-beta-methyl-galactoside. D-Glucose, D-mannose, 2-deoxyglucose, and alpha-methyl-D-glucoside also inhibited lactose uptake. Phosphoenolpyruvate-dependent hydrolysis of o-nitrophenyl-beta-D-galactoside and lactose-dependent release of pyruvate from phosphoenolpyruvate by benzene-treated CT-1 cells showed that CT-1 transports lactose by a phosphoenolpyruvate:sugar phosphotransferase system. Correlations between the growth rate of CT-1 on lactose and properties of the transport system indicated that transport is the rate limiting step in utilization of lactose.
Properties of a mutant Escherichia coli phosphoenolpyruvate carboxylase deficient in coregulation by intermediary metabolites
Authors: McAlister L E , Evans E L , Smith T E
Published: January 2020

Phosphoenolpyruvate carboxylase of Escherichia coli is activated by three different mechanisms: contiguous by acetyl coenzyme A, precursor by fructose 1,6-bisphosphate, and compensatory feedback by cytidine 5'-diphosphate (CDP). Even though each activator can interact independently with the enzyme, synergistic effects are observed with some combinations, namely, fructose 1,6-bisphosphate or CDP (coregulators), with acetyl coenzyme A. A mutant was isolated that has a phosphoenolpyruvate carboxylase which is refractory to activation by fructose, 1,6-bisphosphate and CDP. The mutant enzyme was shown to be active primarily as the dimer and to lack cooperativity in substrate binding. The binding of acetyl coenzyme A and substrate, however, was essentially the same as that of the wild-type enzyme. The mutant cells grew extremely slowly on glucose alone as the sole carbon source. The only defect in the mutant appeared to be the inability of this enzyme to be activated by the coregulators. These data are consistent with the thesis that coregulation by fructose 1,6-bisphosphate or CDP is an essential requirement for the activation in vivo of this enzyme.
Evolutionary relationships among gamma-carboxymuconolactone decarboxylases
Authors: Yeh W K , Durham D R , Fletcher P , Ornston L N
Published: January 2020

gamma-Carboxymuconolactone decarboxylase (EC 4.1.1.44) from Azotobacter vinelandii resembled the isofunctional enzymes from Acinetobacter calcoaceticus and Pseudomonas putida. All three decarboxylases appeared to be hexamers formed by association of identical subunits of about 13,300 daltons. The A. vinelandii and P. putida decarboxylases cross-reacted immunologically with each other, and the NH2-terminal amino acid sequences of the enzymes differed in no more than 7 of the first 36 residues. In contrast, the A. calcoaceticus decarboxylase did not cross-react with the decarboxylase from A. vinelandii or P. putida; the NH2-terminal amino acid sequences of these enzymes diverged about 50% from the NH2-terminal amino acid sequence of the A. calcoaceticus decarboxylase.
Plasmid deoxyribonucleic acid replication in Streptomyces griseus
Authors: Xue Y , Zhuang Z , Zhu Y , Xu Y , Dong K
Published: January 2020

A series of electron micrographs showing the presence of different molecular forms representing various replication stages of plasmid deoxyribonucleic acid from Streptomyces griseus was obtained. Based upon an analysis of these electron micrographs, a tentative model for plasmid deoxyribonucleic acid replication in S. griseus is proposed.
Cloning and characterization of the Escherichia coli gene coding for alkaline phosphatase
Authors: Berg P E
Published: January 2020

The Escherichia coli structural gene for alkaline phosphatase, phoA, and a promoter-like mutant of phoA, called pho-1003(Bin) phoA+, were cloned by using plasmid vectors. Initially, these genes were cloned on deoxyribonucleic acid fragments of 28.9 kilobases (kb). Subsequently, they were subcloned on fragments and 4.8 and then 2.7 kilobases. A restriction map was developed, and phoA was localized to a 1.7-kb region. The promoter end of the gene was inferred by its proximity to another gene cloned on the same deoxyribonucleic acid fragment, proC. The stability of the largest plasmid (33.3 kb) was found to be recA dependent, although the subcloned plasmids were stable in a recA+ strain. Synthesis of alkaline phosphatase directed by the phoA+ and pho-1003(Bin) phoA+ plasmids in a phoA deletion strain was assayed under repressing and derepressing levels of phosphate. These data were compared with the copy numbers of the plasmids. It was found that synthesis of alkaline phosphatase was tightly regulated, even under derepressing conditions: a copy number of 17 enabled cells to synthesize only about twofold more enzyme than did cells with 1 chromosomal copy of phoA+. Enzyme levels were also compared for cells containing pho-1003(Bin) phoA+ and phoA+.
Molecular Mechanisms of Pyrimidine Dimer Excision in
Saccharomyces cerevisiae
: Incision of Ultraviolet-Irradiated Deoxyribonucleic Acid In Vivo
Authors: Reynolds Richard J. , Friedberg Errol C.
Published: January 2020

A group of genetically related ultraviolet (UV)-sensitive mutants of
Saccharomyces cerevisiae
has been examined in terms of their survival after exposure to UV radiation, their ability to carry out excision repair of pyrimidine dimers as measured by the loss of sites (pyrimidine dimers) sensitive to a dimer-specific enzyme probe, and in terms of their ability to effect incision of their deoxyribonucleic acid (DNA) during post-UV incubation in vivo (as measured by the detection of single-strand breaks in nuclear DNA). In addition to a haploid
RAD
+
strain (S288C), 11 different mutants representing six
RAD
loci (
RAD1, RAD2, RAD3, RAD4, RAD14
, and
RAD18
) were examined. Quantitative analysis of excision repair capacity, as determined by the loss of sites in DNA sensitive to an enzyme preparation from
M. luteus
which is specific for pyrimidine dimers, revealed a profound defect in this parameter in all but three of the strains examined. The
rad14-1
mutant showed reduced but significant residual capacity to remove enzyme-sensitive sites as did the
rad2-4
mutant. The latter was the only one of three different
rad2
alleles examined which was leaky in this respect. The UV-sensitive strain carrying the mutant allele
rad18-1
exhibited normal loss of enzyme-sensitive sites consistent with its assignment to the
RAD6
rather than the
RAD3
epistatic group. All strains having mutant alleles of the
RAD1, RAD2, RAD3, RAD4
, and
RAD14
loci showed no detectable incubation-dependent strand breaks in nuclear DNA after exposure to UV radiation. These experiments suggest that the
RAD1, RAD2, RAD3, RAD4
(and probably
RAD14
) genes are all required for the incision of UV-irradiated DNA during pyrimidine dimer excision in vivo.
Formylation of methionyl-transfer ribonucleic acid in Mycoplasma mycoides subsp. mycoides
Authors: Neale G A , Mitchell A , Finch L R
Published: January 2020

Mycoplasma mycoides subsp. mycoides grows readily but does not formylate methionyl-transfer ribonucleic acid in a defined medium without an added formyl donor. Formylation occurs when the medium is supplemented with N5,N10-methenyltetrahydrofolate or N10-formyltetrahydrofolate, but not with folate.
Thiomethylation of tyrosine transfer ribonucleic acid is associated with initiation of sporulation in Bacillus subtilis: effect of phosphate concentration
Authors: Buu A , Menichi B , Heyman T
Published: January 2020

The thiomethylation of Bacillus subtilis tyrosine transfer ribonucleic acid (tRNATyr) (i6A) has been shown to occur during the slowing-down of growth. The extent of this modification in stationary-phase cells grown in defined medium has been determined in parallel with the sporulation frequency. We observed that the presence of phosphate repressed sporulation and also inhibited the thiomethylation of tRNATyr (i6A) of B. subtilis W168. These effects were partially eliminated by decreasing the glucose concentration until it was growth limiting. In the case of strain W23S, in which sporulation is insensitive to glucose repression, sporulation and tRNATyr thiomethylation were not inhibited by nonlimiting concentrations of phosphate. These results suggest that both sporulation and tRNATyr hyper-modification share some common regulatory process.
Cell wall proteins of Aquaspirillum serpens
Authors: Koval S F , Murray R G
Published: January 2020

The Triton X-100-insoluble wall fraction of Aquaspirillum serpens VHA contained three major proteins: the regularly structured (RS) superficial protein (molecular weight 140,000) and two peptidoglycan-associated proteins (molecular weights, 32,000 and 33,000). The molecular arrangement and interactions of the outer membrane and RS proteins were examined with the use of bifunctional cross-linking reagents. The peptidoglycan-associated and RS proteins were not readily cross-linked in either homo- or heteropolymers. This suggests that the free amino groups are not suitably disposed for cross-linking. Some high-molecular-weight multimers of the RS protein were produced, but the subunit structure of the RS array was not stabilized by cross-linking. The peptidoglycan-associated proteins were cross-linked to high-molecular-weight multimers, but no dimers or trimers were produced. This result suggests that these proteins exist in the outer membrane as multimers larger than trimers.
Transformation of
Azotobacter vinelandii
with Plasmids RP4 (IncP-1 Group) and RSF1010 (IncQ Group)
Authors: David Michel , Tronchet Maurice , Dénarié Jean
Published: January 2020

Multicopy plasmid RSF1010 and four of its in vitro-constructed derivatives were mobilized by the self-transmissible RP4 plasmid into
Azotobacter vinelandii
UW. Modifications of the
Escherichia coli
transformation procedure of Cohen et al. (Proc. Natl. Acad. Sci. U.S.A.
69:
2110–2114, 1972) allowed transformation of
A. vinelandii
strains UW and ATCC 12837 with purified RP4 or RSF1010 deoxyribonucleic acid.
Effect of Long Generation Times on Growth ofBacteroides thetaiotaomicronin Carbohydrate-Limited Continuous Culture
Authors: Kotarski Susan F. , Salyers Abigail A.
Published: January 2020

We investigated the ability ofBacteroides thetaiotaomicron, an obligate anaerobe from human colonic microflora, to grow in a carbohydrate-limited continuous culture at generation times ranging from 3.5 to 28 h per division. Four carbohydrates were tested: glucose,N-acetylglucosamine, glucuronic acid, and glucosamine. At a generation time of 3.5 h per division, the growth yields for bacteria growing on glucose,N-acetylglucosamine, and glucuronic acid were 76, 68, and 50 g of cells (dry weight) per mol of substrate, respectively. Growth yields at 28 h per division were 61, 52, and 37 g/mol of substrate, respectively. When glucosamine was the carbohydrate source, a stable population of bacteria was attainable only at generation times longer than 12 h per division. Growth yields at 15 and 32 h per division were 11 and 33 g/mol of substrate, respectively. There was no significant variation with increasing generation times in the specific activities of selected glycolytic enzymes, of disaccharidases such as α- and β-glucosidases and α- and β-galactosidases, or of the polysaccharidase chondroitin sulfate lyase. By contrast, the pattern of fermentation products varied with both the generation time and the carbon source. At a generation time of 3.5 h per division, the main products from the fermentation of glucose were acetate and succinate, with a trace of propionate. At 28 h per division, propionate concentrations were higher and succinate concentrations were lower than at 3.5 h per division. The products from the fermentation of glucosamine were the same as those from glucose fermentation. However, whenN-acetylglucosamine was fermented, the concentration of acetate was much higher at all generation times than when glucose was the carbon source. When glucuronic acid was the carbon source, acetate was the main fermentation product, and only traces of propionate and succinate were detected. Another characteristic that varied with the growth rate was the ability ofB. thetaiotaomicronto produce the inducible enzyme α-glucosidase when exposed to maltose. The ability of the organism to produce this enzyme declined with increasing generation times.
SP02 particles mediating transduction of a plasmid containing SP02 cohesive ends
Authors: Marrero R , Chiafari F A , Lovett P S
Published: January 2020

SP02 particles that mediate transduction of plasmid pPL1010, a 4.6-megadalton derivative of pUB110 containing an Eco RI endonuclease-generated fragment of SP02 deoxyribonucleic acid that spans the cohesive ends, exhibit three unusual features: the transducing particles have a lower buoyant density than infectious particles; the transduction of pPL1010 occurs at high efficiency; and the transducing activity of the particles is relatively resistant to ultraviolet irradiation when the recipient is recombination proficient. Evidence is presented which indicates that SP02(pPL1010) particles carry the plasmid predominantly as a linear multimer having a molecular mass comparable to that of infectious SP02 deoxyribonucleic acid (ca. 31 megadaltons). The plasmid monomers in the linear multimer appear oriented in the same polarity. The buoyant density difference between infectious and transducing particles appears to be due mainly to the buoyant density difference between pPL1010 (1.699 g/cm3) and SP02 deoxyribonucleic acid (1.702 gm/cm3).
Release of colicin E2 from Escherichia coli
Authors: Pugsley A P , Rosenbusch J P
Published: January 2020

Treatment of Escherichia coli K-12(ColE2.P9) with 500 ng of mitomycin C per ml resulted in rapid and almost synchronous colicin E2 production. Colicin accumulated outside the cytoplasmic membrane, most probably in the periplasmic space. Colicin release occurred during a period in which the turbidity of the culture declined markedly. Periplasmic alkaline phosphatase was released during the same period, but cytoplasmic beta-galactosidase release was delayed.
Transcriptional and post-transcriptional control of ribosomal protein and ribonucleic acid polymerase genes
Authors: Little R , Fiil N P , Dennis P P
Published: January 2020

A partial restriction of ribonucleic acid (RNA) polymerase activity has been used to dissociate the coordinate synthesis of ribosomal proteins and subunits of RNA polymerase and to identify transcriptional and post-transcriptional control signals which regulate the expression of these component genes. Within the beta operon [which has the genetic organization: promoter (p beta), rplJ (L10), r;lL (L7/L12), attenuator, rpoB (beta), rpoC (beta'), terminator], the restriction caused a disproportionate increase between proximal and distal gene transcriptions; the transcriptional intensities of the proximal ribosomal protein genes and the distal RNA polymerase genes were elevated about two- and fourfold, respectively. Transcription within the operon containing four ribosomal protein genes and the RNA polymerase alpha gene was also enhanced, whereas transcription within operons containing only ribosomal protein genes was virtually unaffected by the restriction. It was thus concluded that the mechanisms controlling transcription initiation or attenuation or both in operons containing RNA polymerase subunit genes are coupled to the global rate of RNA synthesis. By introducing the composite ColE1 plasmid pJC701 carrying the proximal portion of the L10 operon, including the beta subunit gene, it was possible to achieve a 10- and a 30-fold range in the transcriptional intensities of the genes specifying L10 and L7/L12 and beta, respectively. Under these conditions, the relative synthesis rates of L7/L12 and beta protein varied by less than 2-fold and by about 15-fold, respectively. These observations corroborate the existence of a post-transcriptional mechanism which severely restricts translation of excess L7/L12 and L10 ribosomal protein messenger RNA; this mechanism is probably important in maintaining the balanced synthesis of ribosome components under conditions in which their messenger RNA levels are dissociated. Furthermore, the observed reduction in the translation efficiency of beta subunit messenger RNA may be related to an inhibitory effect caused by accumulation of RNA polymerase assembly intermediates.
Evolution of alkaline phosphatase in marine species of Vibrio
Authors: Woolkalis M J , Baumann P
Published: January 2020

The evolution of alkaline phosphatase was studied in marine species of Vibrio. Two antisera prepared against purified alkaline phosphatases from Vibrio splendidus and Vibrio harveyi were used to estimate the amino acid sequence divergence of this enzyme in 51 strains belonging to nine species. The methods used were the quantitative microcomplement fixation technique and the Ouchterlony double-diffusion procedure. There was a high degree of congruence between the measurement of the amino acid sequence divergence of alkaline phosphatase and the percentage of deoxyribonucleic acid homology of the different organisms relative to both reference strains (correlation coefficient of -0.89) as well as between the amino acid sequence divergence of alkaline phosphatase and superoxide dismutase (correlation coefficient of 0.92) relative to V. splendidus. These findings supported the view that the evolution of marine species of Vibrio is primarily vertical and that horizontal evolution (involving genetic exchange between species), if significant, is restricted to a minor fraction of the bacterial genome.
Unique lipids in Azotobacter vinelandii cysts: synthesis, distribution, and fate during germination
Authors: Su C J , Sadoff H L
Published: January 2020

Unique lipids found in Azotobacter vinelandii cysts are derived primarily from beta-hydroxybutyrate used to induce encystment. Tracer studies with beta-[14C]hydroxybutyrate showed that the biosynthesis of these compounds during encystment began at 8 to 12 h after induction and reached maximal levels after 2 days, Seventy percent of these unique lipids were found in the central body of the cysts, and 23% were found in the exine. Pyronic compounds, which are located mostly in the central body, were degraded during germination of the cysts, but little change occurred in the phenolic compounds, which are more uniformly distributed in the cysts.
Reduced cadmium transport determined by a resistance plasmid in Staphylococcus aureus
Authors: Tynecka Z , Gos Z , Zajac J
Published: January 2020

The presence of a plasmid harboring a gene for Cd2+ resistance led to markedly reduced Cd2+ uptake via the energy-dependent Mn2+ transport system in Staphylococcus aureus strain 17810R. Cd2+ uptake by the resistant strain via this high-affinity system was seen only at very low Cd2+ concentrations. At high concentrations, Cd2+ was taken up by the resistant strain via a different low-affinity uptake system. Cd2+ uptake via this system was energy dependent but was not blocked by Mn2+. Loss of the plasmid from the resistant strain resulted in Cd2+ sensitivity and unblocking of Cd2+ transport via the Mn2+ carrier in the plasmidless derivative strain 17810S. The energy-dependent Cd2+ uptake by the sensitive strain was inhibited by Mn2+ with kinetics indicating competitive inhibition. It is suggested that the second, low-affinity uptake system for Cd2+ in the resistant strain is the energy-dependent cadmium/proton antiporter, which at low Cd2+ concentrations functions in net Cd2+ efflux.
Identification and characterization of Col plasmids from classical colicin E-producing strains
Authors: Watson R , Rowsome W , Tsao J , Visentin L P
Published: January 2020

A series of transformants have been derived which carry the Col plasmids from 11 E-colicin-producing strains isolated by Fredericq or Hamon. The ColE plasmids identified included four of type E1, five of type E2, and two of a type designated E4 by Horak (Zentralbl. Bakteriol. Parasitenkd. Infektionskr. Hyg. Abt. 1 Orig. Reihe A 233:58-63, 1975). Strain K317 was shown to carry a ColE7 plasmid and a 2.7-megadalton (Md) ColE2imm plasmid which confers immunity to colicin E2 but not the ability to produce colicin. Other plasmids identified in the colicinogenic isolates were a 3.4-Md ColN plasmid and a 1.3-Md Col plasmid of unknown type. The ColE1 plasmids all continued replicating in cultures treated with chloramphenicol. Strains carrying the ColE4 or ColE3-CA38 plasmid exhibited partial sensitivity to their own colicins and exhibited an unusual clearing-zone morphology when overlaid on stabs of colicin E2- or E7-producing strains. Except for ColE1-K53, ColE1-K47, ColE2-CA42, and ColE7-K317, all of the ColE plasmids were found to be about 4.3 Md in size. ColE2-CA42 and ColE7-K317 are both about 3.9 Md and are the only two plasmids of the E2, E3, E4, and E7 types that did not yield a 0.39-Md deoxyribonucleic acid fragment as an EcoRI digestion product. The comparisons of the ColE plasmids suggest several structural and functional relationships among them.
Properties of the Bacillus licheniformis A5 glutamine synthetase purified from cells grown in the presence of ammonia or nitrate
Authors: Donohue T J , Bernlohr R W
Published: January 2020

The glutamine synthetase from Bacillus licheniformis A5 was purified by using a combination of polyethylene glycol precipitation and chromatography on Bio-Gel A 1.5m. The resulting preparation was judged to be homogeneous by the criteria of polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, equilibrium analytical ultracentrifugation, and electron microscopic analysis. The enzyme is a dodecamer with a molecular weight of approximately 616,000, and its subunit molecular weight is 51,000. Under optimal assay conditions (pH 6.6, 37 degrees C) apparent Km values for glutamate, ammonia, and manganese.adenosine 5'-triphosphate (1:1 ratio) were 3.6, 0.4, and 0.9 mM, respectively. Glutamine synthetase activity was inhibited approximately 50% by the addition of 5 mM glutamine, alanine, glycine, serine, alpha-ketoglutarate, carbamyl phosphate, adenosine 5'-diphosphate, or inosine 5'-triphosphate to the standard glutamine synthetase assay system, whereas 5 mM adenosine 5'-monophosphate or pyrophosphate caused approximately 90% inhibition of enzyme activity. Phosphorylribosyl pyrophosphate at 5 mM enhanced activity approximately 60%. We were unable to detect any physical or kinetic differences in the properties of the enzyme when it was purified from cells grown in the presence of ammonia or nitrate as sole nitrogen source. The data indicate that B. licheniformis A5 contains one species of glutamine synthetase whose catalytic activity is not regulated by a covalent modification system.
Coat protein synthesis during sporulation of Bacillus subtilis: immunological detection of soluble precursors to the 12,200-dalton spore coat protein
Authors: Goldman R C , Tipper D J
Published: January 2020

Antibody specific to the 12,200-dalton spore coat protein of Bacillus subtilis was used to detect the synthesis of cross-reacting material during sporulation. Cross-reacting protein was first detected by immunoprecipitation after 4 h of development and represented at least 1 to 2% of the total soluble protein synthesis at 5.5 h. A polypeptide of 21,000 daltons was detected in immunoprecipitates by gel electrophoresis. This polypeptide did not accumulate in sporulating cells and was rapidly turned over at the time of coat deposition. In contrast, a 32,000-dalton polypeptide reacted with antibody when unlabeled cell protein was denatured with sodium dodecyl sulfate, separated by gel electrophoresis, and transferred to nitrocellulose paper. This polypeptide was not detected during cell growth or the first 3.5 h of development but was found to accumulate in sporulating cells at 5.5 h. The lack of detection of this polypeptide by immunoprecipitation of undenatured protein indicates that the antigenic sites which cross-reacted with antibody to the 12,200-dalton protein sequence were not exposed unless the molecular conformation was altered. The 32,000-dalton protein may be a primary translation product which is proteolytically processed into mature spore coat protein via a 21,000-dalton intermediate.
Transcriptional inhibition and production of guanosine polyphosphates in Bacillus subtilis
Authors: Price V L , Brown L R
Published: January 2020

When exponentially growing cells of Bacillus subtilis were treated with rifampin or lipiarmycin, both inhibitors of the initiation of ribonucleic acid synthesis, large amounts of (p)ppGpp accumulated. This accumulation appears to be independent of the ribosome-dependent stringent factor reaction because both relA and relC mutants responded in a manner similar to that of the wild type. The possibility that ribonucleic acid polymerase is directly involved in (p)ppGpp metabolism is discussed.
Partial characterization of lipid A of intraperiplasmically grown Bdellovibrio bacteriovorus
Authors: Nelson D R , Rittenberg S C
Published: January 2020

The lipid A components of substrate cell origin incorporated by Bdellovibrio bacteriovorus during intraperiplasmic growth (D. R. Nelson and S. C. Rittenberg, J. Bacteriol. 147:860-868, 1981) were shown to be integrated into its lipopolysaccharide structure. Lipid A isolated from bdellovibrios grown on Escherichia coli was resolved into two fractions by thin-layer chromatography. Fraction 2 had the same Rf as the single lipid A fraction of axenicaly grown bdellovibrios, and both stained identically with aniline-diphenylamine reagent. Fraction 1 resembled, in Rf and staining reaction, the slower migrating of two lipid A fractions obtained from the E.coli used as the substrate cell. Both fractions 1 and 2 contained glucosamine, a substrate cell-derived compound. Greater than 65% of the fatty acids in fraction 1 were derived from the substrate cell, whereas more than 60% of the fatty acids of fraction 2 were synthesized by the bdellovibrio. Nevertheless, each fraction contained significant amounts of fatty acid of both origins. The substrate cell-derived fatty acids had the same distribution of N-acyl and O-acyl linkages as in E. coli lipid A. The data indicate that the two lipid A moieties in lipopolysaccharide of intraperiplasmically grown bdellovibrios are hybrids of substrate cell-derived and bdellovibrio-synthesized components. The data also suggest that disaccharide units and N- and O-acyl linkages preexisting in the substrate cell lipid A may be conserved. A possible explanation for the unequal distribution of substrate cell-derived material in the two lipid A fractions of the bdellovibrio is suggested.
Analysis of mini-F plasmid replication by transposition mutagenesis
Authors: Bergquist P L , Downard R A , Caughey P A , Gardner R C , Lane H E
Published: January 2020

Derivatives of a mini-F plasmid in which Tn3 is inserted in F deoxyribonucleic acid were obtained, and the sites of insertion for 40 of the derivatives were mapped. Tn3 was found to insert at many sites within mini-F, but most insertions were within the 43.0- to 43.7-kilobase (kb), 44.2- to 44.7-kb, and 45.9- to 46.3-kb segments. Hence, these segments are unnecessary for mini-F replication. Most of the Tn3 derivatives were similar to their parent miniplasmid with respect to copy number, stability, and incompatibility. Insertions at 45.15 kb and near 46.0 kb caused a moderate disruption of copy number control, and insertion at 47.6 kb resulted in unstable maintenance. Deletion derivatives lacking deoxyribonucleic acid between 40.3 and 44.76 kb and between 45.92 and 49.4 kb were obtained. This observation suggests either that mini-F contains a third origin, in addition to those already reported to be at 42.6 and 44.4 kb, or that the reported position of the secondary origin, 44.4 kb, is incorrect and that this origin is between 44.76 and 45.92 kb.
Mutagenesis by insertion of drug resistance transposon Tn7 into a vibrio species
Authors: Thomson J A , Hendson M , Magnes R M
Published: January 2020

A halotolerant, collagenolytic strain of Vibrio sp. was conjugated with an Escherichia coli strain carrying plasmid RP4. The plasmid was transferred to and maintained in the Vibrio and could be subsequently transferred in matings to suitably marked stains of the same species. After conjugation with an E. coli carrying the cointegrate plasmid RP4::Mu cts61::Tn7, Vibrio transconjugants were selected that carried Tn7 inserted into the bacterial chromosome. A large proportion of these transconjugants were auxotrophic, showing that plasmid suicide by Mu can be used to isolate Tn7-derived mutants in Vibrio. Approximately half of the auxotrophs isolate Tn7-derived mutants in Vibrio. Approximately half of the auxotrophs isolated were ilv mutants, all of which exhibited the same phenotype. Thus, although Tn7 insertion can induce auxotrophy, including trp, thy, his and ura, in Vibrio, there does appear to be a hot spot for integration in the ilv operon.
The gene for ribosomal protein L31, rpmE, is located at 88.5 minutes on the Escherichia coli chromosomal linkage map
Authors: Dabbs E R
Published: January 2020

Two mutations resulting in an alteration in large-subunit ribosomal protein L31 were mapped at around 88.5 min on the Escherichia coli chromosomal linkage map. They were located between metB and argH and cotransduced over 90% with metB. These mutations were shown to define the structural gene of protein L31, rpmE.
Genetic mapping of the Salmonella typhimurium pncB locus
Authors: Foster J W , Holley E A
Published: January 2020

The nicotinic acid phosphoribosyltransferase locus pncB was located on the Salmonella typhimurium linkage map counterclockwise relative to pyrC. P22 and P1 transductional analyses revealed linkage of pncB with aroA and pyrD, indicating a pncB map position of approximately 20 map units. The results of these cotransduction experiments also indicated that the genetic map distance between gal and pyrD is greater than the published 2.2 map units.