bluuebunny/crossref_metadata_2025 huggingface dataset:
Ammonium assimilation in Rhizobium meliloti
Authors: Osburne M S , Signer E R
Published: January 2020

We have characterized a mutant of Rhizobium meliloti strain 2011 which cannot use ammonium as a nitrogen source. This mutant, RTm2620, was found to have significantly altered glutamate synthase activity. Both the mutant and the wild-type strains had glutamate dehydrogenase activity, which, although stimulated in the presence of glutamate and ammonium, was apparently insufficient to allow ammonium assimmilation. We conclude that the glutamine synthetase-glutamate synthase pathway may be the normal mode of ammonium assimilation by this strain in the free-living state. Independent revertants of Rm2620 were isolated and fell into two classes. Class I revertants regained partial glutamate synthase activity and had the same levels of glutamate dehydrogenase activity as Rm2620. Class II revertants retained the altered glutamate synthase activity but acquired a very high level of assimilatory glutamate dehydrogenase activity. Both classes were found to be altered in their symbiotic properties, although the original Rm2620 mutant was normal in this regard.
Determination of transcriptional units and gene products from the ftsA region of Escherichia coli
Authors: Lutkenhaus J F , Wu H C
Published: January 2020

Lambda transducing phage gamma 16-2 carries the genes envA, ftsZ, ftsA, ddl, and murC and directs the synthesis of six unique proteins in ultraviolet-irradiated cells. Various derivatives of gamma 16-2 carrying smaller segments of the bacterial deoxyribonucleic acid have also been analyzed for their capacity to direct protein synthesis in ultraviolet-irradiated cells. These results, in combination with genetic results, have allowed the gene product of each of these genes to be assigned. In addition, an unidentified gene was located counterclockwise to murC between murC and murF. Analysis of the direction of transcription indicates that murC, ddl, ftsA, and ftsZ are transcribed clockwise on the Escherichia coli genetic map, and envA is transcribed counterclockwise. In addition, it is shown that each of the genes envA, ftsZ, and ftsA can be expressed independently.
High-frequency conjugal transfer of a gonococcal penicillinase plasmid
Authors: Biswas G D , Blackman E Y , Sparling P F
Published: January 2020

Gonococci containing a 24 X 10(6)-dalton conjugal plasmid were able to mobilize for transfer a smaller, non-self-transmissible penicillinase (Pcr) plasmid with high frequency under appropriate conditions. In some strains, over 10% of donor colony-forming units transferred the Pcr plasmid in a mating of less than 2 h, which suggests that the conjugal system was naturally derepressed. Colony-opacity variants containing different quantities of an approximately 28,000-dalton outer membrane protein were altered in their ability to act as conjugal donors and recipients. Maximal transfer of the Pcr plasmid was observed between transparent donors and recipients, lacking appreciable amounts of the 28,000-dalton protein. Under conditions of high-frequency Pcr plasmid mobilization, no conjugal mobilization of chromosomal markers could be discerned.
Transcription of exogenous and endogenous deoxyribonucleic acid templates in cold-shocked Bacillus subtilis
Authors: Kuhl S J , Brown L R
Published: January 2020

Ribonucleic acid (RNA) synthesis was examined in cold-shocked Bacillus subtilis cells. The cells were grown to mid-log stage, harvested, and cold shocked. RNA synthesis was monitored by the incorporation of [3H]uridine triphosphate or [alpha 32P]adenosine triphosphate into trichloroacetic acid-precipitable material in the presence of all four nucleoside triphosphates. The inhibition of RNA synthesis in cold-shocked cells by lipiarmycin, ethidium bromide, rifampin. or streptolydigin was analyzed using mutant or wild-type cells. Also examined were the effects of temperature, salt concentration, and the addition of polyamines or highly phosphorylated nucleotides. In ultraviolet-irradiated and cold-shocked cells, RNA wynthesis decreased to low levels. The addition of exogenous phi 29 or TSP-1 template to these cells caused a 13- to 20-fold increase in RNA synthesis, as monitored by trichloroacetic acid-precipitable counts. RNA synthesized in the presence of phi 29 deoxyribonucleic acid (DNA) hybridizes mainly to EcoRI fragments A and C of phi 29 DBA, These two fragments direct transcription by purified RNA polymerase in vitro and hybridize to early phi 29 DNA produced in vivo. Our results with TSP-1 DNA in this system indicated that the RNA produced hybridizes to the same fragments as early RNA produced in vivo. Plasmic pUB110 DNA was not transcribed in this system.
Pleiotropic mutations rendering Escherichia coli K-12 resistant to bacteriophage TP1
Authors: Wandersman C , Moreno F , Schwartz M
Published: January 2020

tpo mutations, located at 74 min on the genetic map, rendered Escherichia coli K-12 resistant to TP1, a phage which can use either the OmpF protein or the LamB protein as its receptor. tpo mutants synthesized decreased amounts of OmpF and LamB proteins but increased amounts of the OmpC product, another outer membrane protein. The effect of the tpo mutations in lam B gene expression was transcriptional. It is one facet of the following effect on the maltose regulon: strong decreases in the syntheses of the LamB protein and the periplasmic MalE protein occurred when the regulon was uninduced; a lesser decrease occurred in the syntheses of the LamB protein the MalE protein, and the cytoplasmic MalQ protein (amylomaltase) when the regulon was induced. The tpo mutants were found to be phenotypically identical to the perA mutant recently described by Wanner et al. (J. Bacteriol. 140:229--239, 1979) and to some of the ompB mutants described by Verhoef et al. (Mol. Gen. Genet. 169:137--146, 1979). Mapping and complementation analysis suggested that these three types of mutations belong to the same cistron. Our results bring to at least four the number of clearly distinct phenotypes which can result from mutations at, or close to, ompB, a locus which appears increasingly complex.
Proline: an essential intermediate in arginine degradation in Saccharomyces cerevisiae
Authors: Brandriss M C , Magasanik B
Published: January 2020

Results of studies on proline-nonutilizing (Put-) mutants of the yeast Saccharomyces cerevisiae indicate that proline is an essential intermediate in the degradation of arginine. Put- mutants excreted proline when grown on arginine or ornithine as the sole nitrogen source. Yeast cells contained a single enzyme, delta 1-pyrroline-5-carboxylate (P5C) dehydrogenase, which is essential for the complete degradation of both proline and arginine. The sole inducer of this enzyme was found to be proline. P5C dehydrogenase converted P5C to glutamate, but only when the P5C was derived directly from proline. When the P5C was derived from ornithine, it was first converted to proline by the enzyme P5C reductase. Proline was then converted back to P5C and finally to glutamate by the Put enzymes proline oxidase and P5C dehydrogenase.
Specification of surface mating systems among conjugative drug resistance plasmids in Escherichia coli K-12
Authors: Bradley D E , Taylor D E , Cohen D R
Published: January 2020

Representative plasmids for most incompatibility groups in Escherichia coli K-12 were transferred to a "bald" strain to compare transfer frequencies for liquid and solid media. Standard broth matings were used for a liquid environment, but for solid surface mating, conjugation was allowed to take place on nutrient plates before washing off the cells for transconjugant selection on plates containing appropriate drugs. Plasmids that determine rigid pili transferred at least 2,000x better on plates than in broth. Some plasmids that determine thick flexible pili transferred 45 to 470x better, whereas others transferred equally well in both environments, as did plasmids of the I complex, which determine thin flexible pili. These results clearly distinguished a number of surface mating systems where most plasmids were derepressed for transfer and determined conjugative pili constitutively. The temperature-independent IncH2 plasmid R831b transferred best on plates, but other IncH plasmids transferred equally well in broth. This inconsistency led to the reclassification of R831b as IncM.
Selection of Escherichia coli K-12 chromosomal mutants that prevent expression of F-plasmid functions
Authors: Silverman P , Nat K , McEwen J , Birchman R
Published: January 2020

Chromosomal mutants of Escherichia coli deficient in the expression of F-plasmid functions were selected by mutagenizing F- cells, introducing an F' plasmid into the mutagenized cells by conjugation, and identifying transconjugants resistant to the donor-specific bacteriophage Q beta by a simple spray test. All but 1 of 25 mutants were defective in an extracellular stage of Q beta infection, suggesting that they fail to elaborate F-pili. At least six of these were also deficient as deoxyribonucleic acid donors. More than half of the mutants appear to be altered in peviously undetected chromosomal genes required for the expression of F-related cellular functions.
Antibiotic-tolerant mutants of Streptococcus pneumoniae that are not deficient in autolytic activity
Authors: Williamson R , Tomasz A
Published: January 2020

Several mutants of Streptococcus pneumoniae were isolated that appeared tolerant, to varying extents, to the lytic and bactericidal effects of some antibiotics that inhibit peptidoglycan synthesis, but were not deficient in autolytic activity. The method used to select the mutants was based on the survival of tolerant mutants during treatment with either bacitracin, benzylpenicillin, D-cycloserine plus beta-chloro-D-alanine, or vancomycin. Most (60 to 80%) of the surviving isolates were found to be deficient in autolytic activity, and these were rejected. The smaller proportion that had wild-type sensitivity to deoxycholate-induced lysis was studied further with respect to tolerance to the other antibiotics used in the selection procedures. Two of these mutants (selected by treatment with benzylpenicillin) were tolerant to either benzylpenicillin or D-cycloserine plus beta-chloro-D-alanine, but were supersusceptible, in terms of initiation of lysis, to either bacitracin or vancomycin. The minimal inhibitory concentration values of several antibiotics for these two mutants were identical to those for the wild-type strain. Moreover, the interaction of radioactive benzylpenicillin with the penicillin-binding proteins, examined in whole organisms, also appeared the same as previously found for either wild-type or autolytic-deficient strains of S. pneumoniae.
Purification and partial characterization of neuraminidase from type III group B streptococci
Authors: Milligan T W , Mattingly S J , Straus D C
Published: January 2020

Extracellular neuraminidase from a type III fresh clinical isolate of a group B streptococcus was purified by a combination of salt fractionation, affinity chromatography of Affi-Gel blue, ion-exchange chromatography on diethylaminoethylcellulose, and gel filtration on Sephacryl S-200. These procedures yielded enzyme which was purified approximately 1,000-fold compared with the enzyme found in the original supernatant fluid. This type III streptococcal neuraminidase had a molecular weight of approximately 125,000 as estimated by filtration on Sephacryl S-200 and approximately 106,000 when analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In contrast to the majority of other bacterial neuraminidases, the type III group B streptococcal enzyme had no effect on colominic acid or N-acetylneuramin-lactose; however, it was quite active on bovine submaxillary mucin.
Bacteriophages of methanotrophic bacteria
Authors: Tyutikov F M , Bespalova I A , Rebentish B A , Aleksandrushkina N N , Krivisky A S
Published: January 2020

Bacteriophages of methanotrophic bacteria have been found in 16 out of 88 studied samples (underground waters, pond water, soil, gas and oil installation waters, fermentor cultural fluids, bacterial paste, and rumen of cattle) taken in different geographic zones of the Soviet Union. Altogether, 23 phage strains were isolated: 10 strains that specifically lysed only Methylosinus sporium strains, 2 strains that each lysed 1 of 5 Methylosinus trichosporium strains studied, and 11 strains that lysed Flavobacterium gasotypicum and, at the same time, 1 M. sporium strain. By fine structure, the phages were divided into two types (with very short or long noncontractile tails); by host range and serological properties, they fell into three types. One-step growth characteristics of the phages differed only slightly; the latent period varied from 6 to 8 h, the rise period varied from 4 to 6 h, and the average burst size was 100. All phages had guanine- and cytosine-rich double-stranded deoxyribonucleic acid consisting of common nitrogen bases. The molecular mass of the deoxyribonucleic acid as determined by restriction endonuclease analysis was 29.4 X 10(6) for M. sporium phages and 44 X 10(6) for F. gasotypicum phages. By all of the above-mentioned properties, all phages within each of the groups were completely identical to one another, but differed from phages of other groups. Bacteriophages lysing M. sporium and M. trichosporium GB2 were identical to phages M1 and M4, respectively, which were isolated earlier in the German Democratic Republic on the same methanotrophic species.
Characterization of binding sites for a bacteriocin produced by Mycobacterium smegmatis
Authors: Watanabe T , Saito H
Published: January 2020

A bacteriocin from Mycobacterium smegmatis ATCC 14468 which adsorbed to the cell wall-enriched fraction and the crude lipopolysaccharide preparation from Mycobacterium diernhoferi ATCC 19340 was inhibited by D-glucose and alpha-linked glucosylated derivatives.
Hydroxyamino acid utilization and alpha-ketobutyrate toxicity in Pseudomonas cepacia
Authors: Wong H C , Allenza P , Lessie T G
Published: January 2020

Growth of Pseudomonas cepacia 249 on D-threonine required a mutation to permit D-hydroxyamino acid deaminase formation and L-valine to overcome alpha-ketobutyrate toxicity. Strain 249 lacked a second D-hydroxyamino acid deaminase formed by other strains.
Characterization of Tn904 insertions in octopine Ti plasmid mutants of Agrobacterium tumefaciens
Authors: Ooms G , Klapwijk P M , Poulis J A , Schilperoort R A
Published: January 2020

Seven Tn904 insertion mutants of pTi Ach5 affecting Agrobacterium tumefaciens virulence were studied. The mutant character was shown to be plasmid borne. Four of these mutants were avirulent and carried an insertion in restriction endonuclease HpaI fragment 12, a 3.3-megadalton fragment, which therefore appears to be a Ti plasmid region essential for virulence. Two mutants were attenuated in virulence. The inserts mapped close to HpaI fragment 12. One mutant giving rise to small tumors with excessive adventitious root formation on Kalanchoe daigremontiana carried an insertion in the right side of the common sequence in the deoxyribonucleic acid of the Ti plasmid detected in crown gall tumors. The insertion behavior of Tn904 was studied by analyzing 11 independently isolated and randomly chosen mutants. The Tn904 inserts did not affect oncogenicity, tumor morphology, bacterial transfer functions, octopine catabolism functions, or vital parts of the Ti plasmid, such as the origin of replication. Most of the Tn904 inserts were concentrated in a small part of the map. The size of additional deoxyribonucleic acid as a result of Tn904 inserts varied between 5 and 15 megadaltons. In two cases a Ti plasmid was found with two Tn904 insertions at different positions.
Amber mutation affecting the length of Escherichia coli cells
Authors: Martínez-Salas E , Vicente M
Published: January 2020

An amber mutation in a newly found gene (wee) of Escherichia coli has been isolated from strain OV-2, which harbors a temperature-sensitive suppressor. At 42 degrees C cells of the mutant, OV-25, increased in mass and deoxyribonucleic acid content and divided at normal rates, compared with the wild type under the same growth conditions. Total cell length increased under the restrictive conditions, although at a slightly lower rate. Values of mean cell length and cell volume, contrary to what would be expected from the increment in the rate of increase in particles, mass, and deoxyribonucleic acid, became at 42 degrees C smaller than those found in the wild type. A parallel increase in protein content per length and cell density and a loss of viability were found to occur after four generations at the restrictive temperature. The behavior of strain OV-25 in the absence of the wee gene product could be interpreted in terms of either a faulty regulation of the elongation processes or their abnormal coordination with the cell cycle. The genetic location of the wee gene has been found to be at 83.5 min on the E. coli genetic map.
Conservation and variation of nucleotide sequences in Escherichia coli strains isolated from nature
Authors: Harshman L , Riley M
Published: January 2020

A group of Escherichia coli isolates from nature were compared with one another and with laboratory strains of E. coli with respect to size distribution of chromosomal restriction endonuclease fragments and differences in nucleotide sequences in selected small portions of the genomes. The estimated frequency of base substitutions in nucleotide sequences in and near the trp operons of 26 of the 28 E. coli strains examined ranged from 0.008 to 0.066. Nucleotide sequences in or near lambda prophage homologs were significantly more variable than the sequences in or near trp, tnaA, and thyA genes. Thus, the lambda-homologous regions may have a significant horizontal component in their evolutionary histories, having undergone genetic exchange, whereas the trp, tnaA, and thyA regions may have solely vertical evolutionary histories. The relatedness of the E. coli strains in the genetic regions studied indicated that laboratory strains are not more closely related to one other than they are to isolates from nature. The isolates from natural populations did not form groups related either by host taxa or by geographical region of isolation.
Capacity for postreplication repair correlated with transducibility in Rec- mutants of Bacillus subtilis
Authors: Dodson L A , Hadden C T
Published: January 2020

Bacillus subtilis strains deficient in transduction, transformation, or both were examined for the ability to remove pyrimidine dimers and to convert deoxyribonucleic acid newly synthesized after ultraviolet irradiation to high molecular weight. In one strain deficient in both recombination processes, short pieces of deoxyribonucleic acid synthesized after irradiation were not converted to high molecular weight. Two transformable strains deficient in transduction were also deficient in postreplication repair (i.e., joining of newly synthesized DNA fragments), whereas a nontransformable strain that was normal in transduction was proficient in postreplication repair. None of the transformable strains showed deficiencies in repair resynthesis or ligase activity. Our results suggest that some recombinational events may be common to transduction and postreplication repair but not to transformation, emphasizing the difference between these two pathways for genetic exchange.
Mutations in nif genes that cause Klebsiella pneumoniae to be derepressed for nitrogenase synthesis in the presence of ammonium
Authors: MacNeil D , Brill W J
Published: January 2020

Four Nif+ revertants from strains with polar insertions in nifL, were insensitive to ammonium and amino acid repression of nitrogenase synthesis. These strains have mutations located in or near the nifL region. The derepressed phenotype was dominant in a merodiploid containing a nif+ plasmid. These nif regulatory mutations also suppressed the Nif- phenotype of Gln- strains. Thus, regulation by fixed nitrogen (possible via glutamine synthetase) occurs on the nifLA operon but not on the other six nif operons.
Heterogeneity of deoxyribonucleic acid molecules isolated from Mycobacterium smegmatis
Authors: Norgard M V , Imaeda T
Published: January 2020

Bulk chromosomal deoxyribonucleic acids (DNAs) of Mycobacterium smegmatis strains 607+ (wild type) and 607-1 (Strr) and orange-red pigmented variants (OR) were separated into two distinct bands (types 1 and 2) by cesium chloride density gradient centrifugation. Thermal denaturation analyses showed that type 1 and 2 DNA fragments of these strains possessed guanine plus cytosine contents averaging 69.2% and 60.8%, respectively. Type 1 and 2 DNAs from all strains tested were recovered in relatively equal quantities upon isolation and were found to have similar molecular weights (3.0 x 10(7)). Spectrophotometric assay of DNA reassociation showed that homology between any type 1 and 2 DNA fragments was always very low (29 to 33%), even within the same strain. Homologies among type 1 DNAs isolated from any strain were always high (92 to 98%), whereas homologies between type 2 DNA isolated from OR strains and that from their parental strain 607-1 were lower (51 to 55%). Transformation experiments revealed that methionine, leucine, folic acid, and streptomycin markers were found exclusively in type 1 DNA fragments. In addition to the two types of chromosomal DNA, plasmid DNA possessing a molecular weight of about 4 x 10(6) was found in strain 607-1.
Molecular cloning of an erythromycin resistance determinant in streptococci
Authors: Behnke D , Ferretti J J
Published: January 2020

The erythromycin resistance determinant of plasmid pDB102, a derivative of plasmid pSM19035, was cloned into the single HindIII site of the 3.6-megadalton cryptic Streptococcus mutans plasmid pVA318 and introduced into Streptococcus sanguis strain Challis by transformation. Plasmid pDB201, which was isolated from one of the transformants, consisted of the vector plasmid and the 1.15-megadalton HindIII fragment D of pSM19035. HindIII fragment D contained within it one of the two unique "spacer" sequences of pSM19035. Electron micrographs of self-annealed molecules of the recombinant plasmid revealed classical stem-loop structures, and the resistance determinant of pSM19035 appeared as a transposon-like structure. No differences were observed in either the type or the level of erythromycin resistance by pSM19035 or pDB201. The availability of a cloned erythromycin resistance determinant should be useful for future comparative studies of macrolide, lincosamide, and streptogramin B resistance plasmids in streptococci.
Oriented growth of Blastocladiella emersonii in gradients of ionophores and inhibitors
Authors: Harold R L , Harold F M
Published: January 2020

To investigate whether ion currents help to localize growth and development of Blastocladiella emersonii, we grew the organisms in gradients of various ionophores and inhibitors. Gradients were generated by placing into the culture fine glass fibers coated with insoluble inhibitors; in some cases, inhibitors were adsorbed onto beads of ion-exchange resin. Organisms growing in many of these gradients exhibited a striking tendency for the thalli to grow toward the fiber. This proved to be misleading; the cells grew not toward the source of the ionophore but into the unoccupied zone of inhibition adjacent to the fiber. Fibers coated with gramicidin-D induced marked effects on the growth of the rhizoids, which were greatly enlarged and grew toward and onto the fiber. None of the other inhibitors produced such effects, except for beads coated with the proton conductors tetrachlorosalicylanilide and compound 1799. The results suggest that orientation of rhizoid growth results from enhancement of proton flux across the plasma membrane. Growth of the rhizoids was also strongly oriented by gradients of inorganic phosphate and an amino acid mixture; gradients of glucose, K+, Ca2+, and glutamate were ineffective. We propose that a major physiological function of the rhizoid is to transport nutrients to the thallus. Finally, we examined the effects of a series of benzimidazole antitubulins as well as the cytochalasins. These did not orient growth but grossly perturbed the pattern of cellular organization, producing small spherical cells with multiple stunted rhizoids. The findings are interpreted in terms of the interaction of an endogenous transcellular proton current with elements of the cytoskeleton in the determination of form.
Effect of pH on the rate of Candida utilis cell cycle initiation
Authors: Thomas K C
Published: January 2020

The rate of cell cycle initiation (as determined by the rate of bud emergence) in yeast Candida utilis under ammonium-limited phased cultivation was dependent on the pH at which the yeast was grown.
Genetic behavior of the methicillin resistance determinant in Staphylococcus aureus
Authors: Stewart G C , Rosenblum E D
Published: January 2020

The cotransformation frequency of mecC5 with pur-102 using Staphylococcus aureus C5 deoxyribonucleic acid was found to be approximately 45%. However, in cotransduction studies, there was a 15% cotransduction of purine prototrophy and methicillin sensitivity but, in the reciprocal cross, no purine-prototrophic plus Mecr cotransductants were obtained (frequency less than 0.06%). The data support the hypothesis that the mec determinant resides on an inserted deoxyribonucleic acid sequence in S. aureus and that there is no allelic equivalent in sensitive cells.
Chemical and physical differentiation of superoxide dismutases in anaerobes
Authors: Gregory E M , Dapper C H
Published: January 2020

Superoxide dismutase activity in crude or partially purified cell extracts from several species and strains of obligate anaerobe Bacteroides was inhibited instantaneously by NaN3 and was inactivated rapidly upon incubation with H2O2. The extent of NaN3 inhibition varied from 41 to 93%, and the half-life of the enzymatic activity in 5 mM H2O2 ranged from 1.2 to 6.1 min, depending upon the organism tests. When grown in a defined medium containing 59Fe, Bacteroides fragilis (VPI 2393) incorporated radiolabel into a 40,000-molecular-weight NaN3- and H2O2-sensitive superoxide dismutase but did not incorporate 54Mn into that protein under similar growth conditions. The anaerobe Actinomyces naeslundii (VPI 9985) incorporated 54Mn but not 59Fe into a NaN3-insensitive and H2O2-resistant superoxide dismutase. The apparent molecular weight of the superoxide dismutase from this and several other Actinomyces spp. was estimated to be 110,000 to 140,000. Comparison of these data with studies of homogeneous metallosuperoxide dismutases suggests that the Bacteroides spp. studied contain a ferrisuperoxide dismutase, whereas Actinomyces spp. contain a managanisuperoxide dismutase.
Three genes coding for subunits of the membrane sector (F0) of the Escherichia coli adenosine triphosphatase complex
Authors: Downie J A , Cox G B , Langman L , Ash G , Becker M , Gibson F
Published: January 2020

Two mutant unc alleles, unc-469 and unc-476, have been characterized as affecting a previously undescribed gene, designated uncF. The uncF gene is part of the unc operon (with the gene order being uncBFEAGDC), although some uncertainty remains as to the relative order of the uncF and uncE genes. Mutant strains carrying the uncF469 or uncF476 allele lack the 18,000-molecular-weight component of the F0 sector of the adenosine triphosphatase in the cell membrane but retain the dicyclohexylcarbodiimide-binding protein (molecular weight, 8,400). Conversely, strains carrying mutations in the uncE gene lack the dicyclohexylcarbodiimide-binding protein but retain the 18,000-molecular-weight protein in the cell membrane. Strains carrying mutations in the uncB gene have both the 18,000-molecular-weight protein and the dicyclohexylcarbodiimide-binding protein present in the cell membranes. The three proteins of the F0 portion of the adenosine triphosphatase, viz., 24,000, 18,000, and 8,400 molecular weights, became membrane associated after in vitro transcription-translation with plasmid pAN51 as template. Plasmids carrying deletions which affected the UncBFE region were isolated from plasmid pAN51 and characterized genetically. A comparison of the genes that were absent from the various deletion plasmids with the membrane-associated products formed after in vitro transcription-translation indicated that the uncB gene coded for the 24,000-molecular-weight protein and that the gene order was probably uncBFE. A correlation between length of deoxyribonucleic acid, genes present, and their products is presented in relation to plasmid pAN51.
In vivo role of the relA+ gene in regulation of the lac operon
Authors: Primakoff P
Published: January 2020

Under conditions of amino acid limitation, beta-galactosidase was produced at a 70-fold higher rate in a relA+ strain than in an isogenic relA strain of Escherichia coli K-12. Under identical conditions with the relA+ and relA strains carrying various lac promoter mutations, rates of beta-galactosidase synthesis in relA+ (relative to relA) ranged from 26-fold higher (promoter mutant Pr 13) to only 5-fold higher (promoter mutant PrL8uv5). This promoter specificity was independent of strain background and the means of eliciting amino acid limitation. Addition of cyclic AMP to the growth medium altered the relA+/relA difference for beta-galactosidase synthesis from the wild-type lac promoter. The experiments suggest that the relA+/relA difference in lac expression arises primarily at the point of transcription initiation. The results are discussed in relation to recent in vitro data showing a promoter-specific guanosine 5'-diphosphate 3'-diphosphate stimulation of lac transcription (P. Primakoff and S. W. Artz, Proc. Natl. Acad. Sci. U.S.A. 76:1726-1730).
Cell wall turnover in batch and chemostat cultures of Bacillus subtilis
Authors: De Boer W R , Kruyssen F J , Wouters J T
Published: January 2020

Wall turnover was studied in Bacillus subtilis. The loss of radioactively labeled wall polymers was followed during exponential growth in batch and chemostat cultures. Turnover kinetics were identical under all growth conditions; pulse-labeled wall material was lost with first-order kinetics, but only after exponential growth for 1 generation time after its incorporation. Similarly, continuously labeled cells showed an accelerating decrease in wall-bound radioactivity starting immediately after removal of the labeled precursor and also reached first-order kinetics after 1 generation time. A mathematical description was derived for these turnover kinetics, which embraced the concept of "spreading" of old wall chains (H. M. Pooley, J. Bacteriol. 125:1127-1138, 1976). Using this description, we were able to calculate from our experimental data the rate of loss of wall polymers from cells and the fraction of the wall which was sensitive to turnover. We found that about 20% of the wall was lost per generation time and that this loss was affected by turnover activity located in the outer 20 to 45% of the wall; rather large variations were found with both quantities and also between duplicate cultures. These parameters were quite independent of the growth rate (the specific growth rate varied from 1.3 h-1 in broth cultures to 0.2 to 0.3 h-1 in chemostat cultures) and of the nature of the anionic polymer in the wall (which was teichoic acid in cultures with an excess of phosphate and teichuronic acid in phosphate-limited chemostat cultures). Some implications of the observed wall turnover kinetics for models of wall growth in B. subtilis are discussed.
Reducing terminus of O-hapten accumulated in a Salmonella montevideo galE mutant
Authors: Heasley F A
Published: January 2020

The O-haptenic polysaccharide of Salmonella montevideo has been reported to contain glyceraldehyde at its reducing terminus. However, O-hapten preparations from a pmi galE mutant contained products of partial hydrolysis of lipopolysaccharide, which in separate experiments gave [3H]glycerol upon treatment with perchloric acid and [3H]aBH4. Further study of the O-hapten reducing terminus suggested that it was actually mannose.
Physical maps of Klebsiella aerogenes and Salmonella typhimurium hut genes
Authors: Blumenberg M , Magasanik B
Published: January 2020

The recognition sites for several restriction endonucleases were mapped within deoxyribonucleic acid coding for histidine utilization (hut) genes of Salmonella typhimurium and Klebsiella aerogenes. Deoxyribonucleic acid fragments containing the two hut promoters were identified by ribonucleic acid polymerase binding.
Plasmid transfer and genetic recombination by protoplast fusion in staphylococci
Authors: Götz F , Ahrné S , Lindberg M
Published: January 2020

The experimental conditions for plasmid transfer and genetic recombination in Staphylococcus aureus and some coagulase-negative staphylococci by protoplast fusion are described. Protoplasts were prepared by treatment with lysostaphin and lysozyme in a buffered medium with 0.7 to 0.8 M sucrose. Regeneration of cell walls was accomplished on a hypertonic agar medium containing succinate and bovine serum albumin. Transfer of plasmids occurred after treatment of the protoplast mixtures with polyethylene glycol (molecular weight, 6,000) not only between strains of the same species but also between parents of different species, although at approximately 100 times lower frequency in the latter case. Recombination of the chromosomal genes in fused protoplasts required simultaneous treatment of the mixed protoplasts with polyethylene glycol and CaCl2. A method was developed for isolation of recombinants after fusion between mutants of S. areus carrying unselectable markers. Antibiotic resistance plasmids were introduced into the parental strains and used as primary markers to detect protoplast fusion. Chromosomal recombinants were found among the clones with both parental plasmids at a high frequency. The method appears to have simple applications in the construction of strains with multiple mutant characters.
Incorporation of leucine into phospholipids of Bacteroides thetaiotaomicron
Authors: Smith R D , Salyers A A
Published: January 2020

L-[4,5-3H]- or L-[U-14C]leucine was incorporated by Bacteroides thetaiotaomicron into acid-precipitable material even when the bacteria were treated with concentrations of tetracycline high enough to prevent growth. Similar results were obtained when L-[2,3,4-3H]valine or L-[4,5-3H]isoleucine was used instead of leucine. In bacteria which had been treated with tetracycline, the acid-precipitable label was not solubilized by treatment with protease, lysozyme, or deoxyribonuclease. However, virtually all of the label was extractable with chloroform-methanol, indicating that the label had been incorporated into membrane lipids. Since L-[1-14C]leucine was not incorporated into lipids, leucine was probably decarboxylated before incorporation. When a chloroform extract from bacteria which had been labeled with both [32P]phosphate and [3H]leucine was resolved into component phospholipids by two-dimensional thin-layer chromatography, 3H was incorporated into all of the phospholipids. When these phospholipids were deacylated, the 3H from leucine was associated with released fatty acids rather than with the head groups. Thus, it appears that B. thetaiotaomicron can utilize leucine and similar amino acids not only by incorporating them into protein but also by incorporating portions of these amino acids into membrane phospholipids.
Structure of a naturally occurring plasmid with genes for enterotoxin production and drug resistance
Authors: Mazaitis A J , Maas R , Maas W K
Published: January 2020

A physical map of the 117-kilobase conjugative plasmid pCG86 was constructed using electron microscope heteroduplex analysis. This plasmid carries the genes elt, for heat-labile enterotoxin, and estA, for heat-stable enterotoxin, as well as the genes for resistance to tetracycline, streptomycin, sulfonamides, and mercury. These genes were mapped using deletions and Tn5 insertions as physical markers. Analysis of a heteroduplex between pCG86 and a previously described enterotoxin plasmid (EntP307) showed a 48-kilobase region of complete homology which included the genes elt and estA. An 8.8-kilobase BamHI fragment of EntP307 carrying elt, cloned by others, was also shown to be completely homologous with pCG86. The position of elt on the fragment was verified, and it was shown to carry estA as well. A 44-kilobase region of pCG86 showed partial homology with the region of EntP307 previously shown to contain conjugal transfer genes. The gene for tetracycline resistance is carried on a stem-loop structure with the dimensions of Tn10, and the genes for the other drug resistance markers are carried on a 14.6-kilobase segment that forms an insertion loop in heteroduplexes with EntP307. These studies suggest that pCG86 arose either by recombination between an enterotoxin plasmid of incompatibility group FI, like EntP307, and a multiple resistance factor of incompatibility group FII, or by transposition into EntP307 of two transposons.
Identification and preliminary characterization of Vibrio cholerae outer membrane proteins
Authors: Kelley J T , Parker C D
Published: January 2020

Outer membrane proteins of Vibrio cholerae were purified by sucrose density centrifugation and Triton X-100 extraction at 10 mM Mg2+. The proteins were separated by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. V. cholerae outer membrane proteins presented a unique pattern when compared with the patterns of other gram-negative rods. There were 8 to 10 major bands (Mr 94,000 to 27,000), with most of the protein located in band 5 (Mr approximately 45,000), which thus appears to be the major structural protein of the outer membrane. Lipid and carbohydrate were associated with band 6.
Passive rotation of flagella on paralyzed Salmonella typhimurium (mot) mutants by external rotatory driving force
Authors: Ishihara A , Yamaguchi S , Hotani H
Published: January 2020

Salmonella typhimurium mot mutants are unable to rotate their flagella. Dark-field light microscopy showed that the flagella could be rotated passively by an external rotatory driving force.
Selection for loss of tetracycline resistance by Escherichia coli
Authors: Maloy S R , Nunn W D
Published: January 2020

An improved medium for the direct, positive selection of tetracycline-sensitive clones from a population of tetracycline-resistant strains of Escherichia coli is described.
Purification and properties of endo-alpha-1,3-glucanase from a Streptomyces chartreusis strain
Authors: Takehara T , Inoue M , Morioka T , Yokogawa K
Published: January 2020

An enzyme hydrolyzing the water-insoluble glucans produced from sucrose by Streptococcus mutans was purified from the culture concentrate of Streptomyces chartreusis strain F2 by ion-exchange chromatography on diethylaminoethyl cellulose and carboxymethyl cellulose columns and gel filtration on Bio-Gel A-1.5m. The purification achieved was 6.4-fold, with an overall yield of 27.3%. Electrophoresis of the purified enzyme protein gave a single band on a sodium dodecyl sulfate-polyacrylamide gel slab. Its molecular weight was estimated to be approximately 68,000, but there is a possibility that the native enzyme exists in an aggregated form or is an oligomer of the peptide subunits, have a molecular weight larger than 300,000. The pH optimum of the enzyme was 5.5 to 6.0, and its temperature optimum was 55 degrees C. The enzyme lost activity on heating at 65 degrees C for 10 min. The enzyme activity was completely inhibited by the presence of 1 mM Mn2+, Hg2+, Cu2+, Ag2+, or Merthiolate. The Km value for the water-insoluble glucan of S. mutans OMZ176 was an amount of glucan equivalent to 1.54 mM glucose, i.e., 0.89 mM in terms of the alpha-1,3-linked glucose residue. The purified enzyme was specific for glucans containing an alpha-1,3-glucosidic linkage as the major bond. The enzyme hydrolyzed the S. mutans water-insoluble glucans endolytically, and the products were oligosaccharides. These results indicate that the enzyme elaborated by S. chartreusis strain F2 is an endo-alpha-1,3-glucanase (EC 3.2.1.59).
Molybdenum accumulation and storage in Klebsiella pneumoniae and Azotobacter vinelandii
Authors: Pienkos P T , Brill W J
Published: January 2020

In Klebsiella pneumoniae, Mo accumulation appeared to be coregulated with nitrogenase synthesis. O2 and NH+4, which repressed nitrogenase synthesis, also prevented Mo accumulation. In Azotobacter vinelandii, Mo accumulation did not appear to be regulated Mo was accumulated to levels much higher than those seen in K. pneumoniae even when nitrogenase synthesis was repressed. Accumulated Mo was bound mainly to a Mo storage protein, and it could act as a supply for the Mo needed in component I synthesis when extracellular Mo had been exhausted. When A. vinelandii was grown in the presence of WO2-(4) rather than MoO2-(4), it synthesized a W-containing analog of the Mo storage protein. The Mo storage protein was purified from both NH+4 and N2-grown cells of A. vinelandii and found to be a tetramer of two pairs of different subunits binding a minimum of 15 atoms of Mo per tetramer.
Primary structure of the wall peptidoglycan of leprosy-derived corynebacteria
Authors: Janczura E , Leyh-Bouille M , Cocito C , Ghuysen J M
Published: January 2020

The cell walls isolated from axenically grown leprosy-derived corynebacteria were submitted to various chemical and enzymatic degradations. The glycan strands of the wall peptidoglycan are essentially composed of N-acetylglycosaminyl-N-acetylmuramic acid disaccharide units. Small amounts of N-acetylglycosaminyl-N-glycolylmuramic acid (less than 10%) were also detected. The muramic acid residues of adjacent glycan strands are substituted by amidated tetrapeptide units which, in turn, are cross-linked through direct linkages extending between the C-terminal D-alanine residue of one tetrapeptide and the mesodiaminopimelic acid residue of another tetrapeptide. Such a structure is very similar to that of the wall peptidoglycan found in the taxonomically related microorganisms of the Corynebacterium, Mycobacterium, and Nocardia groups.
Isolation and characterization of amber mutations in gene dnaA of escherichia coli K-12
Authors: Schaus N , O'Day K , Peters W , Wright A
Published: January 2020

Amber mutants with defects in the dnaA gene of Escherichia coli K-12 were isolated after localized mutagenesis of the tna-dnaA region of the chromosome. We isolated 36 mutants defective in the initiation of deoxyribonucleic acid replication as determined by their dependence upon integrative suppression by a P2 sig5 prophage. Three of the 36 mutants were shown to contain amber mutations through the use of a temperature-sensitive amber suppressor. These mutations, which mapped between gyrB and tna, were characterized genetically and biochemically as amber mutations in dnaA.
In vivo and in vitro chemotactic methylation in Bacillus subtilis
Authors: Ullah A H , Ordal G W
Published: January 2020

Two doublets of Bacillus subtilis membrane proteins with molecular weights of 69,000 and 71,000 and of 30,000 and 30,800, were labeled by C3H3 transfer in the absence of protein synthesis. In addition, there was intense methylation of several low-molecular-weight substances. Both doublets were missing in a chemotaxis mutant. The equivalent proteins in Escherichia coli and Salmonella typhimurium are believed to be the methyl-accepting chemotaxis proteins. The higher-molecular-weight doublet bands were increased in degree of methylation upon addition of attractant to the bacteria. A methyltransferase from B. subtilis that methylates the wild-type membrane significantly better than the mutant membrane, using S-adenosylmethionine, has been partly purified. The methylated product was alkali labile and is probably a gamma-glutamyl methyl ester, as in E. coli and S. typhimurium. Ca2+ ion inhibited the methyltransferase, with a Ki of about 80 nM. Analysis of the in vitro methylation product showed labeling of the 69,000-dalton methyl-accepting chemotaxis protein and a low-molecular-weight protein, using wild-type membrane. Labeling of the low-molecular-weight protein but not of the 69,000 dalton protein was observed when the mutant membrane was used. The chemotaxis mutant tumbled much longer than the wild type when diluted away from attractant.
Organization and regulation of the ilvGEDA operon in Salmonella typhimurium LT2
Authors: Berg C M , Shaw K J
Published: January 2020

A total of 102 isoleucine- and isoleucine-valine-requiring (ilv) mutants induced by insertion of the transposable element Tn10 have been classified to cistron by growth requirement, cross-feeding behavior, and enzyme assays. The mutations are in a polycistronic operon transcribed in the order ilvGEDA and in a monocistronic operon ilvC. Analysis of distal gene expression in these polar insertion mutants revealed the existence of two constitutive interval promoters, one preceding ilvE and the other preceding ilvD.
Assimilation of single-stranded donor deoxyribonucleic acid fragments by nucleoids of competent cultures of Bacillus subtilis
Authors: van Randen J , Venema G
Published: January 2020

Lysates containing folded chromosomes of competent Bacillus subtilis were prepared. The chromosomes were supercoiled, as indicated by the biphasic response of their sedimentation rates to increasing concentrations of ethidium bromide. Limited incubation of the lysates with increasing concentrations of ribonucleases resulted in a gradual decrease in the sedimentation velocity of the deoxyribonucleic acid (DNA) until finally a constant S value was reached. Incubation with sonicated, 4,5',8-trimethylpsoralen-monoadducted, denatured, homologous donor DNA molecules at 37 degrees C and concomitant irradiation with long-wave ultraviolet light of the nucleoid-containing lysates resulted in the formation of complexes of the donor DNA molecules and the recipient chromosomes. This complex formation was stimulated when nucleoids were previously (i) unfolded by ribonuclease incubation, (ii) (partially) relaxed by X irradiation, or (iii) subjected to both treatments. Monoadducts were not essential. On the other hand, the complex-forming capacity of recipient chromosomes previously cross-linked by 4,5',8-trimethylpsoralen diadducts was greatly reduced, suggesting that strand separation of the recipient molecule was involved in the formation of the complex. None of these effects has been observed when heterologous (Escherichia coli) donor DNA has been used. When the same kind of experiments were carried out at 70 degrees C, donor-recipient DNA complexes were also formed and required strand separation and homology similar to donor-recipient complex formation at 37 degrees C. However, in contrast to what was found at 37 degrees C, unfolding plus relaxation of the nucleoids, as well as the absence of monoadducts in the donor DNA fragments, resulted in a decrease in complex formation. On the basis of these results, we assume that superhelicity can promote the in vitro assimilation of single-stranded donor DNA fragments by nucleoids of competents B. subtilis cells at 70 degrees C, but that at 37 degrees C a different mechanism is involved.
Genetic and physiological characterization of a spontaneous mutant of Escherichia coli B/r with aberrant control of deoxyribonucleic acid replication
Authors: Choung K K , Estiva E , Bremer H
Published: January 2020

Strain TJK16, a low-thymine-requiring thyA deoB derivative of Escherichia coli B/r A, was found to have an increased initiation mass due to a mutation in a gene affecting the control of initiation of deoxyribonucleic acid replication. In contrast to temperature-sensitive initiation mutants, initiation in TJK16 was not temperature sensitive. By phage P1 transduction, it was found that the mutation lies within a small region of the chromosome between dnaA and gyrB; this region includes dnaN and recF. Coumermycin-resistant derivatives of B/r and TJK16 had the same initiation mass as their coumermycin-sensitive parents, and TJK16 had the same sensitivity to coumermycin as the B/r parent, suggesting that the initiation mutation is not in gyrB.