bluuebunny/crossref_metadata_2025 huggingface dataset:
Reduction of iron and synthesis of protoheme by Spirillum itersonii and other organisms
Authors: Dailey H A , Lascelles J
Published: January 2020

Membranes from Spirillum itersonii reduce ferric iron to ferrous iron with reduced nicotinamide adenine dinucleotide or succinate as a source of reductant. Iron reduction was measured spectrophotometrically at 562 nm using ferrozine, which chelates ferrous iron specifically. Reduced nicotinamide adenine dinucleotide or succinate was also effective as a source of iron. The effects of respiratory inhibitors suggested that reduction of iron occurs at one or more sites on the respiratory chain before cytochrome c. Reduction of iron and synthesis of protoheme with the physiological reductants were also observed with crude extracts of other bacteria, including Rhodopseudomonas spheroides, Rhodopseudomonas capsulata, Paracoccus denitrificans, and Escherichia coli. The effect of oxygen upon reduction of iron and formation of protoheme was examined with membranes from S. itersonii, using succinate as a source of reductant. Both systems were inhibited by oxygen, but this effect was completely reversed by addition of antimycin A. We conclude that reduced components of the respiratory chain serve as reductants for ferric iron, but with oxygen present they are oxidized preferentially by the successive members of the chain. This could be a mechanism for regulating synthesis of heme and cytochrome by oxygen.
Induction of D-aldohexoside:cytochrome c oxidoreductase in Agrobacterium tumefaciens
Authors: Nakamura L K , Tyler D D
Published: January 2020

D-Aldohexopyranoside:cytochrome c oxidoreductase (ACO) was strongly induced by cellobiose, alpha-methylglucoside, beta-methylglucoside, kojibiose, and sophorose. Induction was rapid, and ACO was readily detectable within 10 min after addition of cellobiose as inducer. Although not measurable for 30 to 40 min after addition of inducer, once started, the rate of induction with alpha-methylglucoside equaled or even exceeded that obtained with cellobiose. Induction by sucrose, maltose, alpha-alpha-trehalose, melibiose, and lactose was weak. In general, the active ACO inducers were poor glycosidase inducers; the converse also appeared to be true. Although ACO induction was not repressed by D-glucose, it was repressed by succinate, malate, and fumarate.
Energy-linked potassium uptake by mitochondria from wild-type and poky strains of Neurospora crassa
Authors: Smith E W , Slayman C W
Published: January 2020

Mitochondria from Neurospora crassa, like mammalian mitochondria, carry out rapid, energy-linked K+ uptake and H+ release in the presence of valinomycin. The maximal rate of K+ uptake was about 1.0 mumol/mg of mitochondrial protein per min and was seen at valinomycin concentrations in the range of 100 to 200 mug per mg of mitochondrial protein and at K+ concentrations of 4 mM or above. Uptake could be supported either by substrate oxidation or by adenosine 5'-triphosphate (ATP), and was inhibited in the former case by antimycin or cyanide, in the latter case by oligomycin, and in both cases by 2,4-dinitrophenol. Mitochondria from the cytochrome-deficient mutant poky carried out substrate-driven K+ uptake at reduced rates, but oligomycin-sensitive, ATP-driven K+ uptake at rates about 60% greater than those shown by wild-type mitochondria. This result is consistent with the recent finding (Mainzer and Slayman 1976) that poky contains elevated amounts of oligomycin-sensitive mitochondrial adenosine 5'-triphosphatase activity.
Effects of galU mutation on flagellar formation in Escherichia coli
Authors: Komeda Y , Icho T , Iino T
Published: January 2020

Two mutants of Escherichia coli strictly deficient in uridine-diphosphoglucose pyrophosphorylase activity (galU) were found to have very small numbers of flagellar filaments and hooks. In these mutants, both the rate of flagellin (flagellar protein) synthesis and the amount of messenger ribonucleic acid specific for flagellin were considerably lower than in the parental strains. Motile revertants from the galU mutants were isolated and were found to carry a suppressor mutation, which was mapped in the flaH cistron. These strains formed swarms under conditions of catabolite repression; their intracellular concentration of cyclic adenosine 5'-monophosphate was the same as that in the parental strains. These results suggest that the outer membrane affects flagellar formation through the flaH gene product.
Glutamine synthetase of Klebsiella aerogenes: genetic and physiological properties of mutants in the adenylylation system
Authors: Janssen K A , Magasanik B
Published: January 2020

Mutations resulting in defects in the adenylylation system of glutamine synthetase (GS) affect the expression of glnA, the structural gene for GS. Mutants with lesions in glnB are glutamine auxotrophs and contain repressed levels of highly adenylylated GS. Glutamine-independent revertants of the glnB3 mutant have acquired an additional mutation at the glnE site. The glnE54 mutant is incapable of adenylylating GS and produces high levels of enzyme, even when ammonia is present in the growth medium. The fact that mutations in glnB and glnE simultaneously disturb both the normal adenylylation and repression patterns of GS in Klebsiella aerogenes indicates that the adenylylation system, or adenylylation state, of GS is critical for the regulation of synthesis of GS.
Deoxyribonucleic acid repair in vitro by extracts of Escherichia coli
Authors: Masker W E
Published: January 2020

Deoxyribonucleic acid (DNA) from bacteriophage T7 has been used to monitor the capacity of gently lysed extracts of Escherichia coli to perform repair resynthesis after ultraviolet (UV) irradiation. Purified DNA damaged by up to 100 J of UV radiation per m2 was treated with an endonuclease from Micrococcus luteus that introduces single-strand breaks in irradiated DNA. This DNA was then used as a substrate to study repair resynthesis by extracts of E. coli. It was found that incubation with the extract and exogenous nucleoside triphosphates under suitable assay conditions resulted in removal of all pyrimidine dimers and restoration of the substrate DNA to its original molecular weight. Repair resynthesis, detected as nonconservative, UV-stimulated DNA synthesis, was directly proportional tothe number of pyrimidine dimers introduced by radiation. The repair mode described here appears to require DNA polymerase I since it does no occur at the restrictive temperature in polA12 mutants, which contain a thermolabile polymerase. The addition of purified DNA polymerase I to extracts made from a polA mutant restores the ability to complete repair at the restrictive temperature.
NOVEL Escherichia coli dnaB mutant: direct involvement of the dnaB252 gene product in the synthesis of an origin-ribonucleic acid species during initiaion of a round of deoxyribonucleic acid replication
Authors: Zyskind J W , Smith D W
Published: January 2020

The initiation process of deoxyribonucleic acid (DNA) replication in Escherichia coli has been studied using the thermoreversible dna initiation mutant E. coli HfrHl65/120/6 dna-252. This dna mutation was incorrectly classed as a dnaA mutation. Biochemical and genetic evidence suggests that the dna-252 mutant is a novel dnaB mutant, possessing phenotypic properties which distinguish it from other dnaB mutants. Sensitivity of reinitiation in the dna-252 mutant to specific inhibitors of protein, ribonucleic acid (RNA), and DNA synthesis was studied. Reinitiation is shown to be sensitive to rifampin and streptolydigin but not to cholramphenicol. Thus, the dna-252 gene product appears to be required during the initiation process for a step occurring either before or during synthesis of an RNA species (origin-RNA). Using reversible inhibition of RNA synthesis by streptolydigin of a streptolydigin-sensitive derivative of the dna-252 mutant, the dna-252 gene product is shown to be directly involved in the synthesis of an orgin-RNA species. These results are included in a schematic model presented in the accompanying paper of the temporal sequence of events occurring during the initiation process.
Identification of a membrane protein associated with expression of the surface exclusion region of the F transfer operon
Authors: Minkley E G , Ippen-Ihler K
Published: January 2020

Membrane preparations from radioactively labeled male and female strains of Escherichia coli K-12 were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography. An intensely labeled band corresponding to a protein of molecular weight of 24,000 was readily apparent in preparations from Hfr and F-prime strains but not in those from female strains. When preparations from a series of Hfr strains containing transfer operon deletions were examined, presence of the band was found to be associated with retention of the region of the F transfer operon between ilzA and traD. Thus, the band ("protein S") appears to be the product of an F tra operon activity corresponding to traS (the gene for surface or entry exclusion), or an unknown gene in its vicinity. As predicted, protein S was subject to Fin+ control; only a faint band was detectable if the repressed plasmid R100 was also present in the F lac strain. A 24,000-dalton protein was also found in membrane preparations from strains carrying the derepressed plasmids R100-1 and R1-19 but not in those from strains carrying the repressed plasmids R100 or R1. Thus, the appearance of protein S in the membrane may be a general phenomenon resulting from transfer operon expression of F-like plasmids.
Action of a major outer cell envelope membrane protein in conjugation of Escherichia coli K-12
Authors: Schweizer M , Henning U
Published: January 2020

Protein II, a major outer cell envelope membrane protein, was found together with lipopolysaccharide to stoichiometrically inhibit conjugation in Escherichia coli K12.
Formation and dissimilation of oxalacetate and pyruvate Pseudomonas citronellolis grown on noncarbohydrate substrates
Authors: O'Brien R W , Taylor B L
Published: January 2020

Metabolism of lactate as a carbon source by Pseudomonas citronellolis occurred via a nicotinamide adenine dinucleotide (NAD)-independent L-lactate dehydrogenase, which was present in cells grown on DL-lactate but was not present in cells grown on acetate, aspartate, citrate, glucose, glutamate, or malate. The cells also possessed a constitutive, NAD-independent malate dehydrogenase instead of the conventional NAD-dependent malate dehydrogenase instead of the conventional NAD-dependent enzyme in the tricarboxylic acid cycle. Both enzymes were particulate and used dichlorophenolindo-phenol or oxygen as an electron acceptor. In acetate-grown cells, the activity of pyruvate dehydrogenase and NAD phosphate-linked malate enzyme decreased, cells grown on glucose or lactate. This was consistent with the need to maintain a supply of oxalacetate for metabolism of acetate via the tricarboxylic acid cycle. Changes in enzyme activities suggest that gluconeogenesis from noncarbohydrate carbon sources occurs via the malate enzyme (when oxalacetate decarboxylase is inhibited) or a combination of the NAD-independent malate dehydrogenase and oxalacetate decarboxylase.
Phosphoribulokinase from Nitrobacter winogradskyi: activation by reduced nicotinamide adenine dinucleotide and inhibition by pyridoxal phosphate
Authors: Kiesow L A , Lindsley B F , Bless J W
Published: January 2020

CO2 fixation by particle-free extracts from Nitrobacter winogradskyi increased by addition of reduced nicotinamide adenine dinucleotide (NADH). Ribulose-1,5-diphosphate, however, increased CO2 fixation, even in the absence of NADH. Phosphoribulokinase (EC 2.7.1.19) was the enzyme of Nitrobacter extracts that was activated specifically by NADH. Pyridoxal-5-phosphate inhibited both CO2 fixation and NADH-activated phosphoribulokinase from Nitrobacter. However, it did not affect phosphoribulokinase from spinach leaves. Since the spinach enzyme had also no requirement for reduced pyridine nucleotides, it appears that pyridoxal phosphate interferes only with the binding of NADH and not with the binding of ribulose-5-phosphate and adenosine-5'-triphosphate. The regulation of phosphoribulokinase activity by NADH provided Nitrobacter with an energy-dependent control mechanism of CO2 assimilation.
Chemotactic mechanism of Salmonella typhimurium: preliminary mapping and characterization of mutants
Authors: Warrick H M , Taylor B L , Koshland D E
Published: January 2020

Some new, generally nonchemotactic mutants of Salmonella typhimurium were isolated and they, together with previously isolated mutants and some from other investigators, were mapped. Most of the mutants were classified in nine complementation groups, which are probably individual genes. Of these, five map at the end of the flagella region and appear in the order motB-(cheWcheP)-cheX-cheQ-cheR-flaC. Two of the mutations, cheU and cheV, map in the flaQ and flaAII genes, respectively. The remaining genes, cheS and cheT, have not yet been mapped. Most of the mutants are phenotypically smoothly swimming, but some are constantly tumbling. Two of the groups show dominant behavior as recipients in genetic crosses; the rest are recessive. The mutants vary in their responses to stimuli but, since their responses to all chemoeffectors are abnormal, the central processing, rather than individual, receptors must be impaired. The two mutations that coincide with genes for flagella probably involve the locus of the final delivery of sensing signal to the flagella.
Isolation and characterization of a Saccharomyces cerevisiae mutant deficient in pyruvate kinase activity
Authors: Sprague G F
Published: January 2020

A mutant of the yeast Saccharomyces cerevisiae that is deficient in pyruvate kinase activity has been isolated. The mutant strain is capable of growth when supplied with lactate as the carbon source but not capable of growth when supplied with dextrose or other fermentable sugars or glycerol as the carbon source. Genetic analysis demonstrated that the phenotype of the pyruvate kinase-deficient strain was due to a single nuclear mutation, which was designated pyk1, and preliminary genetic mapping experiments located the pyk1 locus on chromosome I, 30 centimorgans from the ade1 locus. Adenine nucleotide levels in the mutant and parental strains were compared when the cells were subjected to various growth and starvation conditions. When carbon supply and energy production were dissociated by supplying the mutant strain with dextrose, adenine nucleotide levels fell dramatically. This result suggests that the initial reactions of glycolysis are not rate limiting, nor are they readily inhibited by feedback controls.
Transport of urea at low concentrations in Chlamydomonas reinhardi
Authors: Williams S K , Hodson R C
Published: January 2020

Urea transport into the unicellular green alga Chlamydomonas reinhardi was investigated to further our understanding of controls operating on urea catabolism in this organism. Transport into cells grown with acetate and deprived of ammonia is a saturable process, mediated by at least two systems operating maximally at different external urea concentrations. The lower concentration system, with an apparent Km for urea of 5.1 micron, was the object of detailed study. Transport of urea from a saturating concentration (57 micron) into ammonia- and acetate-grown cells freshly suspended in ammonia-limited medium was not detected. Upon further culturing in the absence of ammonia, derepression occurred with transport ability, first appearing at about 1 h , reaching a maximum at about 2 h, and maintaining this maximum at least 5 h. In contrast to this, CO2-grown cells became derepressed more slowly, and maximum transport ability was not maintained. Addition of ammonia or methylamine (5 mM) during nitrogen deprivation prevented further increases in transport ability and caused loss of previously acquired transport ability. Cycloheximide (10 microng/ml) had a similar effect. Energy uncouplers or dark, anaerobic conditions depressed transport. By these criteria, transport from low urea concentrations is mediated by a process that requires protein synthesis and activation by cellular energy, and the process has a rapid rate of turnover and of deactivation by ammonia.
Biochemical and regulatory properties of Escherichia coli K-12 hisT mutants
Authors: Bruni C B , Colantuoni V , Sbordone L , Cortese R , Blasi F
Published: January 2020

Escherichia coli K-12 hisT mutants were isolated, and their properties were studied. These mutants are derepressed for the histidine operon, map close to the purF locus at about 49.5 min on the E. coli linkage map, and lack pseudouridylate synthetase activity. The defect in this enzyme leads to the absence of pseudouridines in the anticodon loop of several transfer ribonucleic acid species, as evidenced by the altered elution profile on reversed-phase chromatography and resistance to amino acid analogues. Finally, the hisT mutants studied have a reduced growth rate that appears to be linked to hisT, although it is not known whether it is due to the same mutation. The normal generation time can be restored by supplementing the medium with adenine, uracil, and isoleucine.
Regulation of galactose oxidase synthesis and secretion in Dactylium dendroides: effects of pH and culture density
Authors: Shatzman A R , Kosman D J
Published: January 2020

The effects of pH and growth density on the amount of an extracellular enzyme, galactose oxidase, synthesized by the fungus Dactylium dendroides were studied. Growth at a pH below 6.7 caused a decrease in the ability of the organism to release galactose oxidase. The enzyme retained by these fungal cells was liberated whenever the pH was raised to 7.0. Cycloheximide addition failed to inhibit the appearance of this protein; [3H]leucine added prior to pH adjustment was not incorporated into the released protein, These observations indicate the released protein is not newly synthesized protein. The retained enzyme would be secreted slowly over a 2-day period if the pH was not increased. In addition to regulating protein retention, pH was also shown to be associated with vacuolization, cell volume, culture density, and inhibition of protein synthesis. Cultures maintained at low pH were characterized by a dense growth consisting of highly vacuolated, buoyant, fungal hyphae. Increasing the pH from 6 to 7 caused a decrease in vacuole size. Cells grown at neutral pH maintained a lower density of growth and, based on activity measurements, synthesized 33% more galactose oxidase. Furthermore, cultures grown at pH 6.0 and maintained at a lower cell density produced galactose oxidase at a level similar to that of cells grown at neutral pH. Thus, the elevated density of the cell culture was inhibitory to galactose oxidase synthesis. The observed effects on protein synthesis and release were rather specific for galactose oxidase, since other extracellular proteins appeared in the earliest stages of growth.
Morphology of Ureaplasma urealyticum (T-mycoplasma) organisms and colonies
Authors: Razin S , Masover G K , Palant M , Hayflick L
Published: January 2020

The morphology of Ureaplasm urealyticum in broth cultures was studied by phase-contrast microscopy. Most organisms appeared singly or in pairs. Long filaments and long chains of cocci, common in classical mycoplasma cultures, were not observed. On solid medium, U. urealyticum produced "fried-egg" colonies which developed according to the scheme suggested by Razin and Oliver (J. Gen. Microbiol., 1961) for the morphogenesis of the classical mycoplasma colonies. The formation of the peripheral zone of the colonies followed that of the central zone only when growth conditions were adequate, Hence, the appearance of peripheral zones, and consequently the larger colony size, can be taken as an indicator of improved growth conditions. Incubation in an atmosphere of 100% CO2 resulted in significantly larger colonies than in an atmosphere of N2, O2, or air. CO2 acts as a buffer, keeping the pH at the optimal range for Ureaplasma growth (pH 6.0 to 6.5) in the presence of the ammonia produced from the urea hydrolyzed by the organisms. The addition to the medium of 0.01 M urea together with 0.01 M putrescine enabled better growth than with urea alone. Small amounts of phosphate improved growth in an atmosphere of CO2, apparently fulfilling a nutritional role. Under nitrogen, higher phosphate concentrations were required for good growth, apparently serving as a buffer as well as a nutrient. Sodium chloride and sucrose which had been added to increase the tonicity of the medium inhibited growth above 0.1 M. An increase in the agar concentration above 2% resulted in decreased colony size. Likewise, prolonged drying of the agar plates caused a marked decrease in colony size, mostly affecting the peripheral zone. The addition of both urea and putrescine to the growth medium and incubation in a humidified CO2 atmosphere are recommended for improved growth and formation of fried-egg colonies of U. ureaplyticum on agar. It must be emphasized that these experiments were carried out with a laboratory-adapted strain.
Isolation and characterization of Escherichia coli phase variants and mutants deficient in type 1 pilus production
Authors: Swaney L M , Liu Y P , To C M , To C C , Ippen-Ihler K , Brinton C C
Published: January 2020

Type 1 pili of Escherichia coli are the prototype of the somatic class of pili found on many strains of bacteria. As a first step in the genetic analysis of type 1 piliation, an extensive series of nonpiliated derivatives of E. coli K-12 strain AW405, was characterized to produce attached or free pili when examined in the antiserum or appeared to produce attached or free pili when examined in the electron microscope. The derivatives fell into two classes; phase variants and mutants. Phase variants that formed colonies of two distinctive types, one associated with a predominantly piliated (P+), and the other associated with a nonpiliated (P-) phase, were obtained. Each phase could give rise to the other at a relatively high rate, which was greater in the P- to P+ direction during culture in unshaken liquid medium. In addition, 77 Pil- mutants were selected on the basis of a subtle difference in colonial morphology. The mutants reverted, if at all, at a much lower rate than that of the P- to P+ change. The stability of Pil- derivatives grown in unshaken liquid medium was used as a criterion for distinguishing between phase variants and mutants, Phase variation also effected colonial morphology and chemotactic swarming. These properties did not directly depend upon piliation since Pil- mutants were only slightly altered in colonial form and unaltered in chemotactic swarming. Piliation of Pil+ bacteria was quantitatively affected by growth conditions.
Conjugation-induced lysis of Schizosaccharomyces pombe
Authors: Calleja G B , Yoo B Y , Johnson B F
Published: January 2020

About 15% of the conjugating cells of Schizosaccharomyces pombe were observed to lyse spontaneously during the conjugation process. Lysis occurred at the site of union.
Further characterization of the recipient ability of Escherichia coli K-12 bacteriophage-resistant mutants
Authors: Manning P A , Reeves P
Published: January 2020

We extended the study of Escherichia coli mutants defective in conjugation and showed that the mutants with altered lipopolysaccharide, which are defective as recipients with F-like donors, are also defective with the I-like plasmid R64-11. However, the extent of reduction in recipient ability for I-like donors does not correlate either with the effect on recipient ability for F-like donors or with the degree of alteration to the lipopolysaccharide.
Hydrolysis of lithocholate sulfate by Pseudomonas aeruginosa
Authors: Imperato T J , Wong C G , Chen L J , Bolt R J
Published: January 2020

Pseudomonas aeruginosa was found to be able to hydrolyze bile sulfate. This property was observed when lithocholate sulfate was substituted for the sulfur source in the culture medium. The addition of MgSO4 to the medium inhibited the hydrolysis of the bile sulfate.
Initiation and termination of deoxyribonucleic acid replication in bacteria after a stepwise increase in the velocity of replication
Authors: Bremer H , Young R , Churchward G
Published: January 2020

The theoretical relations between replication, initiation, termination, and deoxyribonucleic acid (DNA) accumulation were derived for experiments in which the length of the time required for the replication of the bacterial chromosome (C period) can be varied. This theory enables one to determine absolute values of the C period from kinetics of DNA accumulation after a "stepup" with thymine-requiring bacteria that are subjected to a sudden increase in the exogenous thymine concentration. Application of this method of data evaluation to an observed step-up experiment with a thy-derivative of Escherichia coli B/r (ATCC 12407) indicated that the theory describes the observed post-step accumulation of DNA accurately within experimental errors. It is also concluded that changes in the replication velocity (C) do not measurably affect the timing of initiation events in a culture.
Lipophilic O-antigens in Rhodospirillum tenue
Authors: Weckesser J , Drews G , Indira R , Mayer H
Published: January 2020

Lipopolysaccharides of eight wild-type strains of the phototrophic bacterium Rhodospirillum tenue have been analyzed. All of the lipopolysaccharides are highly lipophilic. The compositions of preparations obtained by the phenol-water or by the phenol-chloroform-petroleum ether procedure are very similar. The polysaccharide moiety, obtained by mild acid hydrolysis of lipopolysaccharide, consists mainly of aldoheptoses: L-glycero-D-mannoheptose is present in all strains, whereas D-glycero-D-mannoheptose is an additional constituent in some strains. Galactosaminuronic acid and two unknown ninhydrin-positive components were detected in the lipopolysaccharides of six strains. Spermidine and putrescine are present in large amounts in a salt-like linkage in the lipopolysaccharides from three strains. 2-Keto-3-deoxyoctonate forms the linkage between the polysaccharide moiety and lipid A. The lipid A fraction contains all the glucosamine and all the D-arabinose present in the lipopolysaccharide. D-Arabinose is an invariable constituent of the lipid A from the Rhodopseudomonas tenue lipopolysaccharides investigated. The principal fatty acids are beta-hydroxycapric, myristic, and palmitic acids. The isolated R. tenue lipopolysaccharides (O-antigens) react with rabbit antisera prepared against homologous cells. The titers in passive hemagglutination are low, similar to those found with enterobacterial R-lipopolysaccharides. R. tenue O-antigens containing only L-glycero-D-mannoheptose and those containing both the L- and D-epimers of glycero-D-mannoheptose could not be differentiated by serological means.
Isolation and characterization of a polynucleotide phosphorylase from Bacillus amyloliquefaciens
Authors: Erickson R J , Grosch J C
Published: January 2020

Bacillus amyloliquefaciens BaM-2 produces large amounts of extracellular enzymes, and the synthesis of these proteins appears to be dependent upon abnormal ribonucleic acid metabolism. A polynucleotide phosphorylase (nucleoside diphosphate:polynucleotide nucleotidyl transferase) was identified, purified, and characterized from this strain. The purification scheme involved cell disruption, phase partitioning, differential (NH4)2SO4 solubilities, agarose gel filtration, and diethylaminoethyl-Sephadex chromatography. The purified enzyme demonstrated the reactions characteristic of polynucleotide phosphorylase: polymerization, phosphorolysis, and inorganic phosphate exchange with the beta-phosphate of a nucleotide diphosphate. The enzyme was apparently primer independent and required a divalent cation. The reactions for the synthesis of the homopolyribonucleotides, (A)n and (G)n, were optimized with respect to pH and divalent cation concentration. The enzyme is sensitive to inhibition by phosphate ion and heparin and is partially inhibited by rifamycin SV and synthetic polynucleotides.
Stepwise selection of defective nitrogen-fixing phenotypes in Escherichia coli K-12 by dimethyl sulfoxide
Authors: Skotnicki M L , Rolfe B G
Published: January 2020

Storage in dimethyl sulfoxide of Escherichia coli K-12 hybrids carrying Klebsiella pneumoniae nif+ genes can result in selection of a defective nitrogen-fixing phenotype. Dimethyl sulfoxide appears to affect particular inner membrane proteins associated with energy metabolism in E. coli K-12.
Identification of polypeptides necessary for chemotaxis in Escherichia coli
Authors: Silverman M , Simon M
Published: January 2020

Molecular cloning techniques were used to construct Escherichia coli-lambda hybrids that contained many of the genes necessary for flagellar rotation and chemotaxis. The properties of specific hybrids that carried the classical "cheA" and "cheB" loci were examined by genetic complementation and by measuring the capacity of the hybrids to direct the synthesis of specific polypeptides. The results of these tests with lambda hybrids and with a series of deletion mutations derived from the hybrids redefined the "cheA" and "cheB" regions. Six genes were resolved: cheA, cheW, cheX, cheB, cheY, and cheZ. They directed the synthesis of specific polypeptides with the following apparent molecular weights: cheA, 76,000 and 66,000; cheW, 12,000; cheX, 28,000; cheB, 38,000; cheY, 8,000; and cheZ, 24,000. The presence of another gene, cheM, was inferred from the protein synthesis experiments. The cheM gene directed the synthesis of polypeptides with apparent molecular weights of 63,000, 61,000, and 60,000. The synthesis of all of these polypeptides is regulated by the same mechanisms that regulate the synthesis of flagellar-related structural components.
Molecular events associated with induction of arginase in Saccharomyces cerevisiae
Authors: Bossinger J , Cooper T G
Published: January 2020

Arginase, the enzyme responsible for arginine degradation in Saccharomyces cerevisiae, is an inducible protein whose inhibition of ornithine carbamoyl-transferase has been studied extensively. Mutant strains defective in the normal regulation of arginase production have also been isolated. However, in spite of these studies, the macromolecular biosynthetic events involved in production of arginase remain obscure. We have, therefore, studied the requirements of arginase induction. We observed that: (i) 4 min elapsed between the addition of inducer (homoarginine) and the appearance of arginase activity at 30 degrees C; (ii) induction required ribonucleic acid synthesis and a functional rna1 gene product; and (iii) production of arginase-specific synthetic capacity occurred in the absence of protein synthesis but could be expressed only when protein synthesis was not inhibited. Termination of induction by inducer removal, addition of the ribonucleic acid synthesis inhibitor lomofungin, or resuspension of a culture of organisms containing temperature-sensitive rna1 gene products in a medium at 35 degrees C resulted in loss of ability for continued arginase synthesis with half-lives of 5.5, 3.8, and 4.5 min, respectively. These and other recently published data suggest that a variety of inducible or repressible proteins responding rapidly to the environment may be derived from labile synthetic capacities, whereas constitutively produced proteins needed continuously throughout the cell cycle may be derived from synthetic capacities that are significantly more stable.
Ultrastructural surface changes associated with dextran synthesis by Leuconostoc mesenteroides
Authors: Brooker B E
Published: January 2020

When Leuconostoc mesenteroides NCDO 1875 was grown in MRS broth and fixed for electron microscopy in the presence of ruthenium red, the cell wall appeared as a triple-layered structure similar to other, gram-positive bacteria. When such logarithmic-phase cultures were exposed to sucrose, the appearance and growth of a uniform layer of electron-dense material was evident on the surface of the cell wall. After 2 h in the presence of sucrose, the formation of this surface coat (110 to 130 nm thick) was complete. For 85 to 90% of the cells, continued exposure to sucrose did not produce any further change in their appearance, but the rest of the population began to accumulate insoluble capsular dextran at the surface of their coat material. Within 18 h, these cells had produced a large capsule (maximum diameter, 6 micrometer) composed mainly of an extensive reticulum of fine filaments. Periodate-reactive carbohydrate was localized cytochemically in the capsular dextran and in the surface coat of all cells. It is suggested that the surface coat of sucrose-grown cells represents a cell-bound dextran-dextransucrase complex and that the acapsulate cells produce the relatively soluble S dextran reported by previous workers.
Ultrastructural changes during encystment and germination of Bdellovibrio sp
Authors: Tudor J J , Conti S F
Published: January 2020

Under proper conditions, Bdellovibrio sp. strain W cells develop into bdellocysts in appropriate prey bacteria. After attachment and penetration of the prey cell, the encysting bdellovibrio began to accumulate inclusion material and increase in size, and was surrounded by an outer layer of amorphous electrondense material. The cytoplasm of the encysting cell appeared more electron dense, and nuclear areas appeared more compact. During germination of bdellocysts, the outer wall was uniformly broken down the inclusion material changed shape and affinity for the heavy metal stain, and the nuclear areas expanded. As the outer wall was dissolved, outgrowth began with the elongation of the germinant as it emerged from the prey ghost as an actively motile cell.
Mucopeptide Biosynthesis by Minicells of
Escherichia coli
Authors: Reeve John N.
Published: January 2020

Minicells produced by
Escherichia coli
χ925 incorporated amino acids and
N
-acetyl-
d
-glucosamine into mucopeptide.
Phthalate metabolism in Pseudomonas testosteroni: accumulation of 4,5-dihydroxyphthalate by a mutant strain
Authors: Nakazawa T , Hayashi E
Published: January 2020

A mutant strain of Pseudomonas testosteroni blocked in phthalate catabolism converted phthalate into 4,5-dihydroxyphthalate. The latter compound was isolated, and its physical properties were determined. A stoichiometric conversion of the compound to protocatechuate was demonstrated spectrophotometrically with crude extracts of a protocatechuate 4,5-dioxygenase-deficient mutant. Therefore, phthalate is metabolized through 4,5-dihydroxyphthalate and protocatechuate, which is further degraded by protocatechuate 4,5-dioxygenase in P. testosteroni. By using several mutants blocked in phthalate catabolism, 4,5-dihydroxyphthalate decarboxylase was shown to be induced by phthalate. A simple spectrophotometric assay for the enzyme is also reported.
Induction of alkaline phosphatase in Escherichia coli: effect of procaine hydrochloride
Authors: Tribhuwan R C , Pradhan D S
Published: January 2020

The effect of procaine hydrochloride, an anesthetic known to alter membrane structure, on the induced formation of alkaline phosphatase, a periplasmic enzyme, in Escherichia coli was investigated. Procaine hydrochloride specifically arrested the appearance of active alkaline phosphatase while permitting the induction of another enzyme, beta-galactosidase, which is internally localized. Evidence has been obtained to show that procaine hydrochloride does not arrest synthesis of inactive monomer subunits of the enzyme, indicating that the drug interferes in the conversion of monomer subunits to an active dimer enzyme.
Characterization of an endonuclease associated with the drug resistance plasmid pKM101
Authors: Lackey D , Walker G C , Keng T , Linn S
Published: January 2020

An endonuclease was detected in strains of Salmonella typhimurium containing the drug resistance plasmid pKM101. The enzyme was not detectable in strains lacking this plasmid, but it was present in strains containing mutants of pKM101 that were no longer able to enhance host cell mutagenesis. The endonuclease had a molecular weight of roughly 75,000 and, at pH 7.0, was equally active on single-stranded and duplex deoxyribonucleic acid (DNA). The reaction with single-stranded DNA was optimal at pH 5.5, whereas with duplex DNA the optimum was pH 6.8. The enzyme required a divalent cation for activity, and it had no detectable exonuclease activity with single-stranded or duplex DNA. The endonuclease extensively degraded DNA with no apparent base specificity, forming 5'-phosphomonoester termini. Although characterization of the endonuclease has not revealed its function, the enzyme does not appear to be a restriction endonuclease.
Two modes of metabolic regulation of lysyl-transfer ribonucleic acid synthetase in Escherichia coli K-12
Authors: Hirshfield I N , Liu C , Yeh F M
Published: January 2020

Lysyl-transfer ribonucleic acid (tRNA) synthetase activity was compared in three independently isolated Escherichia coli K-12 mutants of the enzyme S-adenosyl-L-methionine synthetase (metK mutants) and their isogenic parents. In all three cases the activity of the lysyl-tRNA synthetase was elevated two- to fourfold in the mutant strains. Glycyl-L-leucine (3 mM) usually enhanced lysyl-tRNA synthetase activity two- to threefold in wild-type cells but did not further stimulate the synthetase activity in metK mutants. By two other criteria, the lysyl-tRNA synthetase from wild-type cells grown with the peptide and from the metK mutant RG62, grown in minimal medium, were similar. These criteria are enhanced resistance to thermal inactivation and altered susceptibility to endogenous proteases when compared with the synthetase from wild-type cells grown in minimal medium. In a separate set of experiments, the activities of the lysyl-, arginyl-, seryl-, and valyl-tRNA synthetases were measured in an isogenic pair of relt and rel strains of E. coli grown in a relatively poor growth medium (acetate) and in enriched medium. In the rel+ strain the level of all four synthetases was higher (two- to fourfold) in the enriched medium as expected. In the rel strain the difference in the activities of the synthetases between the two media were diminished. In all four cases the activities of the synthetases were higher in acetate medium in the rel strain. Evidence is presented that these two modes of metabolic regulation act independently.
Localization of acid phosphatase in Saccharomyces cerevisiae: a clue to cell wall formation
Authors: Linnemans W A , Boer P , Elbers P F
Published: January 2020

Acid phosphatase is present in two layers of the cell envelope of Saccharomyces cerevisiae. These are separated by another layer, which is free of acid phosphatase. We have evidence that the cell wall is built up in two stages, which are independent. In the first stage, the cell wall is built up during the formation of the bud. Glucanase vesicles are involved in this process. In the second stage, a thick layer is deposited at the inside against the new cell wall. This results in the thick, rigid wall of the mature yeast cell. This latter layer is probably assembled on the outer surface of the plasmalemma.
Occurrence of a class II restriction endonuclease in Staphylococcus aureus
Authors: Stobberingh E E , Schiphof R , Sussenbach J S
Published: January 2020

The occurrence of class II restriction endonucleases (enzymes that both recognize and cleave a specific nucleotide sequence in deoxyribonucleic acid [DNA] in Staphylococcus aureus has been investigated by analysis of crude extracts obtained from different propagating strains of the International Phage Typing System. Of the four main groups of strains in the International System, only extracts of group II strains were found to contain class II restriction endonucleases. The identical cleavage patterns obtained by incubation of different DNAs with cell extracts of group II strains suggest that these enzymes all recognize and cleave the same nucleotide sequence. This recognition site has been determined to be 5'-G-A-T-C-3'-3'-C-T-A-G-5' for the prototype of these enzymes, Sau3AI (J. S. Sussenbach et al., Nucleic Acids Res. 3:3192-3202, 1976). Evidence is presented that the classification of group II strains is based on restriction modification and is correlated with the presence of a class II restriction enoduclease that recognizes and cleaves the above sequence.
Antibody to the
d
-Alanine Carboxypeptidase of
Bacillus subtilis
Does Not Cross-React with Other Penicillin-Binding Proteins
Authors: Buchanan Christine E. , Hsia Judy , Strominger Jack L.
Published: January 2020

The fact that antibody to
d
-alanine carboxypeptidase of
Bacillus subtilis
does not cross-react with other penicillin-binding proteins suggests that these proteins are not precursors or multimers of the enzyme.
Nitrogen regulation of arginase in Neurospora crassa
Authors: Vaca G , Mora J
Published: January 2020

The final products of the arginine catabolism that can be utilized as a nitrogen source in Neurospora crassa are ammonium, glutamic acid, and glutamine. The effect of these compounds on arginase induction by arginine was studied. In wild-type strain 74-A, induction by arginine was almost completely repressed by glutamic acid plus ammonium, whereas ammonium or glutamic acid alone had only moderate effects. Arginine products of catabolism also repressed arginase induction. A mutant, ure-1, which lacks urease activity, hyperinduced its arginase with arginine as a nitrogen source. The addition of either ammonium or glutamine produced effects similar to those in the wild-type strain. The effect of ammonium on arginase induction is mediated through its conversion into glutamine. This was demonstrated in mutant am-1, which lacks L-glutamate dehydrogenase activity. In this mutant, the effect of glutamic acid was reduced, and, with ammonium, it was completely lost. The addition of glutamine or glutamic acid plus ammonium to this strain decreased by threefold the induction of arginase by arginine. Proline, a final product of arginine catabolism, competitively inhibited arginase activity. This effect and the repression of arginase by glutamine are examples of negative modulation of the first enzyme in a catabolic pathway by its final products.
Regulation of the Neurospora crassa assimilatory nitrate reductase
Authors: Ketchum P A , Zeeb D D , Owens M S
Published: January 2020

Reduced nicotinamide adenine dinucleotide phosphate (NADPH)-nitrate reductase from Neurospora crassa was purified and found to be stimulated by certain amino acids, citrate, and ethylenediaminetetraacetic acid (EDTA). Stimulation by citrate and the amino acids was dependent upon the prior removal of EDTA from the enzyme preparations, since low quantities of EDTA resulted in maximal stimulation. Removal of EDTA from enzyme preparations by dialysis against Chelex-containing buffer resulted in a loss of nitrate reductase activity. Addition of alanine, arginine, glycine, glutamine, glutamate, histidine, tryptophan, and citrate restored and stimulated nitrate reductase activity from 29- to 46-fold. The amino acids tested altered the Km of NADPH-nitrate reductase for NADPH but did not significantly change that for nitrate. The Km of nitrate reductase for NADPH increased with increasing concentrations of histidine but decreased with increasing concentrations of glutamine. Amino acid modulation of NADPH-nitrate reductase activity is discussed in relation to the conservation of energy (NADPH) by Neurospora when nitrate is the nitrogen source.
Alteration of the Bacillus subtilis glutamine synthetase results in overproduction of the enzyme
Authors: Dean D R , Hoch J A , Aronson A I
Published: January 2020

A mutational leading to glutamine auxotrophy was located near a 5-fluorouracil resistance marker in the citB-thyA region of the Bacillus subtilis chromosome. This mutation resulted in a glutamine synthetase with altered kinetic and feedback properties. The specific activity of manganese-stimulated glutamine synthetase activity in crude extracts was 18-fold higher, and the magnesium-stimulated activity was about 30% that of the wild type. Quantitation of the enzyme by precipitation with antibody prepared against pure enzyme confirmed the presence of high enzyme levels in the mutant. This mutation is very closely linked (recombination index of 0.03) to another glutamine auxotroph containing enzyme with altered electrophoretic and heat sensitivity properties. Mutations in the structural gene for glutamine synthetase may result not only in altered catalytic and regulatory properties but also in altered production of the enzyme.
Ribonucleic Acid Polymerase Mutant of
Escherichia coli
Defective in Flagella Formation
Authors: Yamamori T. , Ito K. , Yura T. , Suzuki T. , Iino T.
Published: January 2020

Escherichia coli
K-12 mutants that are resistant to bacteriophage χ, defective in motility, and unable to grow at high temperature (42°C) were isolated from among those selected for rifampin resistance at low temperature (30°C) after mutagenesis with
N
-methyl-
N
′-nitro-
N
-nitrosoguanidine. Genetic analysis of one such mutant indicated the presence of two mutations that probably affect the β subunit of ribonucleic acid (RNA) polymerase: one (
rif
) causing rifampin resistance and the other (Ts-74) conferring resistance to phage χ (and loss of motility) and temperature sensitivity for growth. Observations with an electron microscope revealed that the number of flagella per mutant cell was significantly reduced, suggesting that the Ts-74 mutation somehow affected flagella formation at the permissive temperature. When a mutant culture was transferred from 30 to 42°C, deoxyribonucleic acid synthesis accelerated normally, but RNA or protein synthesis was enhanced relatively little. The rate of synthesis of β and β′ subunits of RNA polymerase was low even at 30°C and was further reduced at 42°C, in contrast to the parental wild-type strain. Expression of the lactose and other sugar fermentation operons, as well as lysogenization with phage λ, occurred normally at 30°C, suggesting that the mutation does not cause general shut-off of gene expression regulated by cyclic adenosine 3′,5′-monophosphate.
Transformation in Escherichia coli: cryogenic preservation of competent cells
Authors: Morrison D A
Published: January 2020

Escherichia coli cells prepared for transformation by treatment with cold 0.1 M CaCl2 remained viable and competent after storage at -82 degrees C in 15% glycerol; thawed-cell samples yielded up to 10(6) transformants per microgram of plasmid deoxyribonucleic acid.
Lysozyme-promoted association of protein I molecules in the outer membrane of Escherichia coli
Authors: Chopra I , Howe G B , Ball P R
Published: January 2020

Incubation of whole envelopes prepared from sonically oscillated Escherichia coli K-12 cultures with lysozyme in vitro resulted in the appearance of a protein species with an apparent molecular weight double that of outer membrane protein I. Similar dimers were also detected in purified outer membranes and whole envelopes from lysozyme-induced spheroplasts of E. coli K-12. This was confirmed by two-dimensional electrophoresis in which the dimers were resolved in the second dimension to run as single polypeptides of protein I. Formation of dimers was correlated with peptidoglycan degradation, but the ability of protein I molecules to associate may vary between strains of E. coli, since dimers were found only in outer membranes from E. coli W7. We suggest that extensive degradation of peptidoglycan leads to nonspecific formation of protein I aggregates, but that these aggregates do not occur in vivo.