bluuebunny/crossref_metadata_2025 huggingface dataset:
Selective plasmid transduction in Bacillus pumilus
Authors: Bramucci M G , Lovett P S
Published: January 2020

The inducible temperate bacteriophage phi75 and a clear-plaque-forming variant, phi75C1, mediated transduction of a 4.4 X 10(6)-dalton multicopy Bacillus pumilus plasmid, pPL10, at frequencies of 10(-5) to 10(-6) transductants per plaque-forming unit. phi75- and phi75C1-mediated transduction of several chromosome markers tested did not occur at a detectable frequency. phi75-mediated plasmid transducing activity resides in particles that are similar to infectious particles in sedimentation velocity and buoyant density.
Component from the cell surface of the hydrocarbon-utilizing yeast Candida tropicalis with possible relation to hydrocarbon transport
Authors: Käppeli O , Fiechter A
Published: January 2020

A polysaccharide-fatty acid complex was isolated from the cell surface of Candida tropicalis growing on alkanes. This complex was solubilized by Pronase treatment of whole cells. A decrease in alkane-binding affinity was observed after Pronase treatment, resulting in 10 to 12% of the yeast dry cell weight being released as polysaccharide. The isolated polysaccharide contained 2.5% fatty acids. C. tropicalis and Saccharomyces cerevisiae grown with glucose contained only traces of fatty acids in the corresponding polysaccharide fraction. The fatty acids were not removed from the polysaccharide moiety by gel filtration. Extraction of the polysaccharide with chloroform-methanol showed that fatty acids were covalently bound to the polysaccharide. The amphipathic nature of the isolated polysaccharide and the hydrocarbon-induced formation suggest a possible role in alkane metabolism.
Hydrogenase Activity and the H
2
-Fumarate Electron Transport System in
Bacteroides fragilis
Authors: Harris Martha A. , Reddy C. Adinaryayana
Published: January 2020

Hydrogenase activity and the H
2
-fumarate electron transport system in a carbohydrate-fermenting obligate anaerobe,
Bacteroides fragilis
, were investigated. In both whole cells and cell extracts, hydrogenase activity was demonstrated with methylene blue, benzyl viologen, flavin mononucleotide, or flavin adenine dinucleotide as the electron acceptor. A catalytic quantity of benzyl viologen or ferredoxin from
Clostridium pasteurianum
was required to reduce nicotinamide adenine dinucleotide or nicotinamide adenine dinucleotide phosphate with H
2
. Much of the hydrogenase activity appeared to be associated with the soluble fraction of the cell. Fumarate reduction to succinate by H
2
was demonstrable in cell extracts only in the presence of a catalytic quantity of benzyl viologen, flavin mononucleotide, flavin adenine dinucleotide, or ferredoxin from
C. pasteurianum
. Sulfhydryl compounds were not required for fumarate reduction by H
2
, but mercaptoethanol and dithiothreitol appeared to stimulate this activity by 59 and 61%, respectively. Inhibition of fumarate reduction by acriflavin, rotenone, 2-heptyl-4-hydroxyquinoline-
N
-oxide, and antimycin A suggest the involvement of a flavoprotein, a quinone, and cytochrome
b
in the reduction of fumarate to succinate. The involvement of a quinone in fumarate reduction is also apparent from the inhibition of fumarate reduction by H
2
when cell extracts were irradiated with ultraviolet light. Based on the evidence obtained, a possible scheme for the flow of electrons from H
2
to fumarate in
B. fragilis
is proposed.
Mercury and Organomercurial Resistances Determined by Plasmids in
Pseudomonas
Authors: Clark Dan L. , Weiss Alison A. , Silver Simon
Published: January 2020

Mercury and organomercurial resistance determined by genes on ten
Pseudomonas aeruginosa
plasmids and one
Pseudomonas putida
plasmid have been studied with regard to the range of substrates and the range of inducers. The plasmidless strains were sensitive to growth inhibition by Hg
2+
and did not volatilize Hg
0
from Hg
2+
. A strain with plasmid RP1 (which does not confer resistance to Hg
2+
) similarly did not volatilize mercury. All 10 plasmids determine mercury resistance by way of an inducible enzyme system. Hg
2+
was reduced to Hg
0
, which is insoluble in water and rapidly volatilizes from the growth medium. Plasmids pMG1, pMG2, R26, R933, R93-1, and pVS1 in
P. aeruginosa
and MER in
P. putida
conferred resistance to and the ability to volatilize mercury from Hg
2+
, but strains with these plasmids were sensitive to and could not volatilize mercury from the organomercurials methylmercury, ethylmercury, phenylmercury, and thimerosal. These plasmids, in addition, conferred resistance to the organomercurials merbromin,
p
-hydroxymercuribenzoate, and fluorescein mercuric acetate. The other plasmids, FP2, R38, R3108, and pVS2, determined resistance to and decomposition of a range of organomercurials, including methylmercury, ethylmercury, phenylmercury, and thimerosal. These plasmids also conferred resistance to the organomercurials merbromin,
p
-hydroxymercuribenzoate, and fluorescein mercuric acetate by a mechanism not involving degradation. In all cases, organomercurial decomposition and mercury volatilization were induced by exposure to Hg
2+
or organomercurials. The plasmids differed in the relative efficacy of inducers. Hg
2+
resistance with strains that are organomercurial sensitive appeared to be induced preferentially by Hg
2+
and only poorly by organomercurials to which the cells are sensitive. However, the organomercurials
p
-hydroxymercuribenzoate, merbromin, and fluorescein mercuric acetate were strong gratuitous inducers but not substrates for the Hg
2+
volatilization system. With strains resistant to phenylmercury and thimerosal, these organomercurials were both inducers and substrates.
Genetic locus (ompB) affecting a major outer-membrane protein in Escherichia coli K-12
Authors: Sarma V , Reeves P
Published: January 2020

Three multiply colicin-tolerant mutants in Escherichia coli K-12 from the TolIV, TolXIV, and TolXV phenotypic groups, all lacking or having only trace amounts of protein 1, a major outer-membrane protein, were mapped by Hfr crosses, and the position on the chromosome was confirmed by cotransduction with nearby markers. The mutations were located near malQP in the 74-min region of the E. coli chromosome. This locus is designated ompB, and analysis of data from two three-point crosses determined the linear sequence of genes to be aroB-ompB-malQP-glpD.
2-Hydroxypyridine Metabolism and Pigment Formation in Three
Arthrobacter
Species
Authors: Kolenbrander P. E. , Weinberger M.
Published: January 2020

Three species of the genus
Arthrobacter, A. crystallopoietes, A. pyridinolis,
and
A. viridescens
, have the capabilities to utilize 2-hydroxypyridine (2-HP) as the sole source of carbon and nitrogen for growth and to produce an extracellular crystalline pigment from this substrate. Degradation of 2-HP by cell-free extracts requires the presence of both reduced nicotinamide adenine dinucleotide and molecular oxygen and is stimulated by flavin mononucleotide, suggesting the presence of a monooxygenase activity in the extract. Loss of the ability to produce pigment at a high spontaneous frequency, 0.26% loss per generation, is observed only with
A. crystallopoietes
and can be visualized by the presence of sectored and fully nonpigmented colonies on solid media containing 2-HP. Concomitant with the loss of pigment-producing character are both loss of ability to utilize 2-HP for growth and oxidation of 2-HP by cell-free extracts. These three 2-HP-associated characteristics also are lost simultaneously by treating cultures of
A. crystallopoietes
with curing agents, such as acridine orange and mitomycin C, but are not curable in
A. pyridinolis
or
A. viridescens
. All nonpigmented strains of
A. crystallopoietes
are nonrevertible for these properties. These data suggest that 2-HP-related characteristics are plasmid determined in
A. crystallopoietes
but not in
A. pyridinolis
and
A. viridescens
. A survey for the presence of plasmids in these three species and two physiologically unrelated species,
A. globiformis
and
A. atrocyaneus
, revealed plasmid material only in
A. globiformis
and
A. crystallopoietes
.
Adaptive changes in phosphate uptake by the fungus Neurospora crassa in response to phosphate supply
Authors: Beever R E , Burns D J
Published: January 2020

The phosphate uptake rate of Neurospora crassa germlings growing exponentially in media containing phosphate at concentrations between 10 mM and 50 micronM was virtually constant. The uptake characteristics of these germlings were studied in detail assuming the simultaneous operation of two uptake systems, one of low affinity and one of high affinity. The Km of the low-affinity system was constant after growth at phosphate concentrations greater than 1 mM but became progressively lower as the concentration was reduced below 1 mM. In contrast, the Km of the high-affinity system was independent of the phosphate concentration of the growth medium. The Vmax of each system was highest after growth at low phosphate concentrations. As the phosphate concentration was increased to a maximum of 100 mM, the Vmax of the low-affinity system fell gradually, whereas that of the high-affinity system at first fell rapidly but then reached a constant minimum value at concentrations of 2.5 mM and higher. The differences in the kinetic parameters fully account for the constancy of uptake rate shown by the germlings.
Further mapping of several membrane lipid biosynthetic genes (fabC, fabB, gpsA, plsB) of Escherichia coli
Authors: Clark D , Cronan J E
Published: January 2020

New genetic mapping data on several loci involved in membrane lipid synthesis are reported. We demonstrated that the lesion designated fabC by Broekman and Hoeckstra (Mol. Gen. Genet. 124:65-67, 1975) is a fabB mutation. In the course of this work, the orientation of the pdxB and fabB loci was determined. The order of the loci is fabB pdxB purF. We also report cotransduction between the gpsA and cysE loci and show that the order of these markers is mtl cysE gpsA. Cotransduction between the plsB and kdgK loci was also sought. Despite extensive experiments, we were unable to detect cotransduction between these loci. In addition, we were unable to detect cotransduction among several markers in region 46 to 48 min of the map.
Nitrogen and Ammonia Assimilation in the Cyanobacteria: Purification of Glutamine Synthetase from
Anabaena
sp. Strain CA
Authors: Stacey Gary , Tabita F. Robert , Baalen Chase Van
Published: January 2020

Glutamine synthetase was purified from the cyanobacterium
Anabaena
sp. strain CA, a newly isolated marine organism. This organism grows rapidly under nitrogen-fixing conditions and therefore is ideally suited for studies concerning cyanobacterial nitrogen metabolism. Studies were conducted to optimize the production of glutamine synthetase by
Anabaena
CA. The highest specific activities were obtained from cells grown in the presence of atmospheric N
2
or KNO
3
(13 mM); when NH
4
Cl was used as the nitrogen source, the specific activity was reduced by approximately 40%. Furthermore, through the use of a whole-cell γ-glutamylhydroxamate transferase assay, it was found that the maximum number of enzyme units is obtained in the late logarithmic stage of growth. Glutamine synthetase purification requires only three steps and results in a preparation that is electrophoretically homogeneous. The transferase specific activity (units per milligram of protein) of the purified enzyme is 78, whereas the biosynthetic specific activity is 2.2. The molecular weight of the native protein was found to be approximately 590,000, and the subunit molecular weight was determined to be about 50,000. Thus, this cyanobacterial enzyme closely resembles the enzyme obtained from other procaryotic sources, at least with regard to size. The purification of glutamine synthetase from
Anabaena
CA should stimulate a more detailed study of this enzyme and its role in cyanobacterial nitrogen metabolism.
Genetics of Nitrofurazone Resistance in
Escherichia coli
Authors: McCalla D. R. , Kaiser Christel , Green M. H. L.
Published: January 2020

Wild-type
Escherichia coli
cells are sensitive to nitrofurazone (NF) and many other nitrofuran derivatives. A variety of evidence indicated that these compounds are converted to toxic “active” metabolites by reductases present in the bacteria. Sensitive
E. coli
K-12 acquired threefold-greater resistance to NF in one mutational step. These partially resistant mutants could undergo a second mutation that made them 10 times as resistant as the wild type. Mutation of wild-type strain K-12 to the higher level of resistance in a single step was not observed. The first mutational step was associated with partial loss of reduced nicotinamide adenine dinucleotide phosphate-linked, O
2
-insensitive NF reductase activity, and the second step was associated with loss of the remaining activity. The two-step mutants did, however, contain other NF reductases that were inhibited by O
2
and reduced NF only under anaerobic conditions. We designated the genes that control reductase activity “nitrofuran sensitivity genes” (
nfsA
and
nfsB
). Thus, wild-type strains are
nfsA
+
nfsB
+
, and the resistant double mutants are
nfsA nfsB
. A variety of crosses established that these genes are both located close to
gal
, that the most probable sequence is
lac nfsB gal nfsA
, and that the single-step mutants with an intermediate level of resistance are
nfsA nfsB
+
. The
nfsA
+
nfsB
strains contained about 70 to 80% of the wild-type reductase I activity—apparently enough to confer wild-type sensitivity. This reductase activity was resistant to 2 M urea. The
nfsA nfsB
+
strains had only 20 to 30% of the wild-type activity, and this residual activity was sensitive to 2 M urea.
Transport of antibiotics and metabolite analogs by systems under cyclic AMP control: positive selection of Salmonella typhimurium cya and crp mutants
Authors: Alper M D , Ames B N
Published: January 2020

Mutants in the cyclic AMP (cAMP) control system in Salmonella typhimurium (cya = adenyl cyclase, crp = cAMP receptor protein) were partially resistant to growth inhibition by 22 antibiotics (including fosfomycin, nalidixic acid, and streptomycin) and 29 inhibitory analogs of normal bacterial fuel/carbon sources. This resistance was used as the basis for an efficient positive selection of cya and crp mutants. We propose that these antibiotics and analogs enter the bacteria through transport systems normally used for transporting fuel/carbon sources and that this is accomplished because of a structural similarity between the antibiotic and the natural substrate of the particular transport system involved. We propose that these transport systems are all under positive control by cAMP and that cAMP acts as a signal molecule (alarmone) for fuel/carbon deprivation. Evidence is provided for a hierarchy within operons controlled by cAMP. The methodology is shown to be useful for analyzing both antibiotic transport systems and the cAMP super-control system.
Evolution of Pseudomonas R-plasmids: consequences of Tn1 insertion and resultant partial diploidy to chromosome and Tra- R-plasmid mobilization
Authors: Olsen R H
Published: January 2020

Tn1 transposes from pRO161, a Tra- derivative of RP1, to Pseudomonas aeruginosa sex factor FP2. The acquisition of Tn1 by FP2 results in its ability to mobilize pRO161 to other bacteria. Genetic evidence presented here suggests two sequential mechanisms. Initially, transposition of Tn1 results in trans-diploidy for the Tra+ and Tra- plasmids. This subsequently allows mobilization of the Tra- R-plasmid dependent on a host recombination mechanism. Transconjugants from this mating contain either stable cointegrate R-plasmids or aggregates resulting from dissociation of the cointegrates into a Tra+ and Tra- plasmid. These aggregates have lost at least part of Tn1 from their parent FP2:Tn1 component, but now they mobilize the tra- R-plasmid from a recombination-deficient (Rec-) genetic background as well as from Rec+ donor strains. Transconjugants from these retransfer matings are aggregates. These results suggest a contribution of transposons to R-plasmid evolution and dissemination beyond the mere acquisition of resistance to a given antibiotic.
Low-temperature conditional cell division mutants of Escherichia coli
Authors: Sturgeon J A , Ingram L O
Published: January 2020

Fifteen low-temperature conditional division mutants of Escherichia coli K-12 was isolated. They grew normally at 39 degrees C but formed filaments at 30 degrees C. All exhibited a coordinated burst of cell division when the filaments were shifted to the permissive temperature (39 degrees C). None of the various agents that stimulate cell division in other mutant systems (salt, sucrose, ethanol, and chloramphenicol) was very effective in restoring colony-forming ability at 25 degrees C or in stimulating cell division in broth. One of these mutants, strain JS10, was found to have an altered cell envelope as evidenced by increased sensitivity to deoxycholate and antibiotics, as well as leakage of ribonulcease I, a periplasmic enzyme. This mutant had normal rates of DNA synthesis, RNA synthesis, and phospholipid synthesis at both the nonpermissive and permissive temperatures. However, strain JS10 required new protein synthesis in the apparent absence of new RNA synthesis for division of filaments at the permissive temperature. The division of lesion in strain JS10 is cotransducible with malA, aroB, and glpD and maps within min 72 to 75 on the E. coli chromosome.
Genetics of the alpha-ketoglutarate dehydrogenase complex of Bacillus subtilis
Authors: Hoch J A , Coukoulis H J
Published: January 2020

The enzymatic defects in a number of Bacillus subtilis mutants of the alpha-ketoglutarate dehydrogenase complex lacking activity have been investigated. Mutants in the citK locus, as well as a series of deletions of unknown length covering the citK locus, are deficient in E1 of the complex, alpha-ketoglutarate dehydrogenase, but have normal activities of E2, dehydrolipoyl transsuccinylase, and E3, lipoamide dehydrogenase. The citK mutants and the citL22 mutant show in vitro complementation of alpha-ketoglutarate dehydrogenase complex activity. The citL22 mutant is severely deficient in lipoamide dehydrogenase activity, and, as a result, lacks activity for both the alpha-ketoglutarate and the pyruvate dehydrogenase complexes. Thus, the E3 components of both complexes are identical. The citL22 mutation maps between ura and metC on the chromosome.
Uptake of extracellular biotin by Escherichia coli biotin prototrophs
Authors: Cicmanec J F , Lichstein H C
Published: January 2020

Uptake of exogenous biotin by two Escherichia coli biotin prototroph strains, K-12 and Crookes, appeared to involve incorporation at a fixed number of binding sites located at the cell membrane. Incorporation was characterized as a binding process specific for biotin, not requiring energy, and stimulated by acidic pH. Constant saturation quantities of exogenous biotin were incorporated by these cells, and the amounts, which were titrated, depended on whether the cells were resting or dividing. Resting cells incorporated exogenous biotin amounting to 2% of their total intracellular biotin content. Fifty percent of the exogenous biotin was incorporated into their free biotin fraction, and 50% was incorporated into their bound biotin fraction. On the other hand, dividing cells incorporated exogenous biotin into all of their intracellular sites, 88% going into the intracellular-bound biotin fraction, and 12% going into the free biotin fraction. Calculations suggested that each cell contained approximately 3,000 binding sites for biotin. It was postulated that biotin incorporation sites might have been components of acetyl coenzyme A carboxylase located at or near the membrane.
Preferential inhibitory action of sodium cholate on an Escherichia coli strain carrying a plasmid in an integrated state
Authors: Yoshida Y , Takamatsu N , Yoshikawa M
Published: January 2020

Sodium cholate was shown to be preferentially more active on Escherichia coli strains carrying an integrated plasmid, i.e., on Hfr strains, than on their parental strains with or without a plasmid in an autonomous state.
Comparative studies of porphyrin production in Propionibacterium acnes and Propionibacterium granulosum
Authors: Lee W L , Shalita A R , Poh-Fitzpatrick M B
Published: January 2020

Porphyrin production by Propionibacterium acnes and that by Propionibacterium granulosum were compared. Porphyrin synthesized by both organisms was identified as coproporphyrin III on the bases of absorption and fluorescence spectra and behavior on paper chromatography and thin-layer chromatography. Quantitative, rather than qualitative, differences in production were found between these organisms. In general, P. granulosum produced significantly greater amounts (P less than 0.001) of porphyrin than did P. acnes. delta-Aminolevulinic acid synthetase appeared to be the rate-limiting enzyme of the heme biosynthetic pathway in both organisms. The increased porphyrin production in P. granulosum is apparently associated with increased delta-aminolevulinic acid synthetase activity.
Expression of the uvrB gene of Escherichia coli: in vitro construction of a pMB9 uvrB plasmid
Authors: Pannekoek H , Noordermeer I A , van Sluis C A , van de Putte P
Published: January 2020

Bacteriophage lambdab2att2 [lambdab2cI857intam6(deltabioAB)bioFCD+uvrB+phr+] codes for a function(s) that confers UV resistance (Uvr+) and reactivation of irradiated phage (Hcr+) to an Uvr-Hcr-Escherichia coli strain. It was demonstrated that these functions are expressed under the control of bacterial regulatory elements located on lambdab2att2 DNA. The location of the E. coli uvrB gene on the DNA of this transducing phage was established by heteroduplex and restriction-enzyme analyses. Recombinant DNA molecules were constructed in vitro from plasmid pMB9 (Tcr), as the vector, and an EcoRI fragment (Eco-RI-F) of lambdab2att2 DNA. The resulting plasmid, designated pNP5, has a molecular weight of 5.1 X 10(6) and replicates in a relaxed fashion. Transformation of E. coli uvrB with plasmid pNP5 resulted in clones that are Uvr+ Tcr. Irradiation of bacteria transformed with plasmid pNP5 with low UV doses revealed a complete restoratation of the Uvr+ phenotype by the presence of the cloned EcoRI-F DNA, while only a partial restoration was observed after irradiation with high UV doses. Likewise, the Hcr+ character was also partially restored due to the presence of pNP5. No correlation was found between the acquired Uvr+, Hcr+ properties, and the presence of correndonuclease II activity in an extract of bacteria that harbor plasmid pNP5.
Incomplete flagellar structures in nonflagellate mutants of Salmonella typhimurium
Authors: Suzuki T , Iino T , Horiguchi T , Yamaguchi S
Published: January 2020

Incomplete flagellar structures were detected in osmotically shocked cells or membrane-associated fraction of many nonflagellate mutants of Salmonella typhimurium by electron microscopy. The predominant types of these structures in the mutants were cistron specific. The incomplete basal bodies were detected in flaFI, flaFIV, flaFVIII, and flaFIX mutants, the structure homologous to a basal body in flaFV mutants, the polyhook-basal body complex in flaR mutants, and the hook-basal body complex in flaL and flaU mutants. No structures homologous to flagellar bases or their parts were detected in the early-fla group nonflagellate mutants of flaAI, flaAII, flaAIII, flaB, flaC, flaD, flaE, flaFII, flaFIII, flaFVI, flaFVII, flaFX, flaK, and flaM. From these observations, a process of flagellar morphogenesis was postulated. The functions of the early-fla group are essential to the formation of S ring-M ring-rod complexes bound to the membrane. The completion of basal bodies requires succeeding functions of flaFI, flaFIV, flaFVIII, and flaFIX. Next, the formation of hooks attached to basal bodies proceeds by the function of flaFV and by flaR, which controls the hook length. Flagellar filaments appear at the tips of hooks because of the functions of flaL, flaU, and flagellin genes.
Role of glutathione in reversing the deleterious effects of a thiol-oxidizing agent in Escherichia coli
Authors: Hibberd K A , Berget P B , Warner H R , Fuchs J A
Published: January 2020

Diamide was found to be much less specific for the oxidation of glutathione in vivo in Escherichia coli than had been previously assumed. In vivo, only a slight alteration of the ratio of reduced to oxidized glutathione was found, whereas a significant amount of glutathione was found in the form of mixed disulfide with proteins. This latter occurrence was postulated as being responsible for the bacteriostatic effect of diamide.
Outer Membrane Proteins of
Escherichia coli
VII. Evidence That Bacteriophage-Directed Protein 2 Functions as a Pore
Authors: Pugsley Anthony P. , Schnaitman Carl A.
Published: January 2020

Protein 1, a major protein of the outer membrane of
Escherichia coli
, has been shown to be the pore allowing the passage of small hydrophilic solutes across the outer membrane. In
E. coli
K-12 protein 1 consists of two subspecies, 1a and 1b, whereas in
E. coli
B it consists of a single species which has an electrophoretic mobility similar to that of 1a. K-12 strains mutant at the
ompB
locus lack both proteins 1a and 1b and exhibit multiple transport defects, resistance to toxic metal ions, and tolerance to a number of colicins. Mutation at the
tolF
locus results in the loss of 1a, in less severe transport defects, and more limited colicin tolerance. Mutation at the
par
locus causes the loss of protein 1b, but no transport defects or colicin tolerance. Lysogeny of
E. coli
by phage PA-2 results in the production of a new major protein, protein 2. Lysogeny of K-12
ompB
mutants resulted in dramatic reversal of the transport defects and restoration of the sensitivity to colicins E2 and E3 but not to other colicins. This was shown to be due to the production of protein 2, since lysogeny by phage mutants lacking the ability to elicit protein 2 production did not show this effect. Thus, protein 2 can function as an effective pore.
ompB
mutations in
E. coli
B also resulted in loss of protein 1 and similar multiple transport defects, but these were only partially reversed by phage lysogeny and the resulting production of protein 2. When the
ompB
region from
E. coli
B was moved by transduction into an
E. coli
K-12 background, only small amounts of proteins 1a and 1b were found in the outer membrane. These results indicate that genes governing the synthesis of outer membrane proteins may not function interchangeably between K-12 and B strains, indicating differences in regulation or biosynthesis of these proteins between these strains.
Pyridoxine-requiring mutants of Escherichia coli: glycolaldehyde dehydrogenase is not coded for by the pdxB gene
Authors: Shimizu S , Dempsey W B
Published: January 2020

Twenty-seven independent pyridoxineless mutants belonging to genetic linkage group I were assayed for glycolaldehyde dehydrogenase. Some mutants lacked enzyme activity entirely, and others showed activity ranging from very low to wild-type levels. Reversion to pyridoxine independence usually had no effect upon this activity. Transfer of the pyridoxine genes to a common host that had wild-type levels of enzyme activity made the recipient pyridoxineless without affecting the activity. These results negate the idea of an obligatory role for glycolaldehyde dehydrogenase in pyridoxine biosynthesis.
Analysis of cell division in single clones of the Escherichia coli K-12 lexA mutant
Authors: Howe W E , Mount D W
Published: January 2020

The growth of clones of lexA mutant and lexA+ cells was analyzed. During normal growth lexA mutant clones frequently divided early, producing smaller newborn cells than the lexA+ clones. Some newborn cells in the lexA clones did not elongate or divide at all, a response that was never observed in the lexA+ clones. When starved for thymidine, most of the lexA mutant clones elongated and subsequently divided. The majority of lexA+ clones also elongated but did not divide. The above results suggest that one of the functions of the lexA+ gene is coordination of DNA repair with cell division.
Replication of colicin E1 plasmid DNA in vivo requires no plasmid-encoded proteins
Authors: Donoghue D J , Sharp P A
Published: January 2020

A derivative of bacteriophage lambda containing a colicin E1 plasmid replicon was constructed by recombinant DNA techniques. This phage, lambdacol100, has two functional modes of DNA replication; it can replicate via either plasmid or phage replication systems. lambdacol100 has been used to introduce the colicin E1 plasmid replicon into Escherichia coli previously treated with chloramphenicol to block protein synthesis. Under these conditions, lambdacol100 DNA is replicated normally as a colicin E1 plasmid. This suggests that colicin E1 plasmid replication in vivo does not require any plasmid-encoded proteins.
Chitin Synthesis in
Candida albicans
: Comparison of Yeast and Hyphal Forms
Authors: Braun Phyllis Cellini , Calderone R. A.
Published: January 2020

Chitin synthesis was studied in both yeast and hyphae of the dimorphic fungus
Candida albicans
. Incorporation of
N
-acetyl-
d
-[1-
3
H]glucosamine ([
3
H]GluNAc) into an acid-alkali-insoluble fraction was 10 times greater in hyphal-phase cells. A crude preparation of chitin synthetase was obtained from sonically treated protoplasts of both forms of
Candida
. Enzyme activity, which was determined by using [
14
C]UDP-GLuNAc as a substrate, was exclusively associated with the 80,000 ×
g
pellet from sonically treated protoplasts of both forms. It was determined that enzyme activity (nanomoles of [
14
C]UDP-GluNAc incorporated per milligram of protein) was approximately 2 times greater in hyphae versus yeast cells. Enzyme activity in both yeast and hyphae increased six- to sevenfold when the enzyme preparations were preincubated with trypsin. A vacuolar fraction, obtained from yeast cells but not from hyphae, stimulated enzyme activity when incubated with either yeast or hyphal enzyme preparations. Membrane fractions from protoplasts coated with [
3
H]concanavalin A before disruption were isolated by Renografin density gradient centrifugation. Chitin synthetase activity was preferentially associated with the concanavalin A-labeled fraction, suggesting that the enzyme was located on the plasma membrane. In addition, enzyme activity in protoplasts treated with cold glutaraldehyde before disruption was significantly greater than in protoplasts that were sonically disrupted and then treated with cold glutaraldehyde, indicating that the enzyme resides on the inner side of the plasma membrane.
Double Mutants of
Bacillus subtilis
Growing as Helices
Authors: Rogers H. J. , Thurman P. F.
Published: January 2020

Double mutants of
Bacillus subtilis
, deficient in autolysin and
rod
in genotype, grow as helical filaments of unseparated cells when changed from the cocci to rods.
Changes in cyclic AMP levels during development in Myxococcus xanthus
Authors: Yajko D M , Zusman D R
Published: January 2020

Cyclic AMP levels doubled in Myxococcus xanthus under conditions in which cells aggregate and form fruiting bodies. In liquid medium, glycerol- or dimethyl sulfoxide-induced sporulating cultures exhibited a sharp but transient rise in cyclic AMP concentration after 45 min.
Characterization and regulation of anthranilate synthetase from a chloramphenicol-producing streptomycete
Authors: Francis M M , Vining L C , Westlake D W
Published: January 2020

In Streptomyces sp. 3022a, anthranilate synthetase is composed of two non-identical subunits. The major subunit (molecular weight, 72,000) converts chorismic acid to anthranilic acid, using ammonia as the source of the amino group. The smaller subunit (molecular weight 28,000 to 29,000) confers on the enzyme the ability to use glutamine instead of ammonia as a substrate. In this study, reactivity with glutamine reached its maximum at pH 7.2 to 7.6, whereas that with ammonia increased linearly through pH 9.0 without reaching a maximum. Activity was increased and stabilized by adding glutamine and magnesium chloride to the buffer system. Both activities of the enzyme were inhibited by anthranilic acid and by tryptophan. Synthesis was repressed by histidine, anthranilic acid, tryptophan, and p-aminobenzoic acid. When activity was repressed by anthranilic acid and by tryptophan, there was a concomitant increase in the activity of arylamine synthetase, an enzyme involved in chloramphenicol production. Stimulating arylamine synthetase, however, did not increase antibiotic synthesis.
Viability and Endogenous Substrates Used During Starvation Survival of
Rhodospirillum rubrum
Authors: Breznak John A. , Potrikus C. J. , Pfennig Norbert , Ensign Jerald C.
Published: January 2020

Cells of
Rhodospirillum rubrum
were grown photoorganotrophically and chemoorganotrophically and then starved for organic carbon and combined nitrogen under four conditions: anaerobically in the light and dark and aerobically in the light and dark. Illumination prolonged viability and suppressed the net degradation of cell material of phototrophically grown cells, but had no effect on chemotrophically grown cells that did not contain bacteriochlorophyll. The half-life survival times of carbohydrate-rich phototrophically grown cells during starvation anaerobically or aerobically in the light were 17 and 14.5 days, respectively. The values for starvation aerobically and anaerobically in the dark were 3 and 0.5 days, respectively. Chemotrophically grown cells had half-life survival times of 3 and 4 days during starvation aerobically in the light and dark, respectively, and 0.8 day during starvation anaerobically in the light or dark. Of all cell constituents examined, carbohydrate was most extensively degraded during starvation, although the rate of degradation was slowest for phototrophically grown cells starved anaerobically in the light. Phototrophically grown cells containing poly-β-hydroxybutyrate as carbon reserve were less able to survive starvation anaerobically in the light than were carbohydrate-rich cells starved under comparable conditions. Light intensity had a significant effect on viability of phototrophically grown cells starving anaerobically. At light intensities of 320 to 650 lx, the half-life survival times were 17 to 24 days. At 2,950 to 10,500 lx, the survival times decreased to 1.5 to 5.5 days. The kinetics of cell death correlated well with the rate of loss of cell mass of starving cells. However, the cause of death could not be attributed to degradation of any specific cell component.
Temperature-dependent variation in the synthesis of streptococcal group-specific carbohydrate. II. Biosynthetic studies in group A and variant strains
Authors: Ayoub E M
Published: January 2020

The biosynthesis of the cell wall polysaccharide and peptidoglycan of group A and A-486-Var streptococci was studied with N-acetyl-[14C]glucosamine, UDP-N-acetyl-[14C]glucosamine, and [14C]glucose. The incorporation of N-acetyl-[14C]-glucosamine into the cell wall four times greater in the A-486-Var cells than in the group A cells. However, the percentage of the total label incorporated into the cell wall polysaccharide at 37 degrees C by the A-486-Var strain was 12%, compared with 66% for the group A cells. When the A-486-Var was grown at 22 degrees C, the proportion of the label incorporated into the cell wall polysaccharide increased to 41%. At 37 degrees C, N-acetyl-[14C]glucosamine was incorporated preferentially into the peptidoglycan of the A-486-Var; almost three times as much of the label was incorporated into the peptidoglycan at 37 degrees C as was incorporated at 22 degrees C. Studies with protoplast membranes of these organisms showed similar differences, with a fourfold greater uptake of UDP-N-acetyl-[14C]glucosamine by the A-486-Var membranes at both incubation temperatures. These studies suggest that a defect in the incorporation of N-acetylglucosamine into the side chain of the polysaccharide is present in the A-486-Var strain at a step following the synthesis of UDP-N-acetylglucosamine. This defect, which may involve the UDP-N-acetylglucosamine transferase, is temperature dependent in the A-486-Var strain.
Pattern of phenazine pigment production by a strain of Pseudomonas aeruginosa
Authors: Kanner D , Gerber N N , Bartha R
Published: January 2020

An atypical strain of Pseudomonas aeruginosa capable of synthesizing three phenazine pigments was isolated. Cultural conditions, under which the strain forms either chlororaphin, oxychlororaphin, or pyocyanine, are described. This broad spectrum of pigment production, as well as some other characteristics, sets this strain apart from previously described chlororaphin producers.
Reduction of nicotinamide adenine dinucleotide by pyruvate:lipoate oxidoreductase in anaerobic, dark-grown Rhodospirillum rubrum mutant C
Authors: Gorrell T E , Uffen R L
Published: January 2020

Cell extracts from fermentatively grown Rhodospirillum rubrum reduced about 80 nmol of nicotinamide adenine dinucleotide (NAD) per mg of protein per min under anaerobic conditions with sodium pyruvate. The reaction was specific for pyruvate and NAD; NAD phosphate was not reduced. Results indicated that pyruvate-linked NAD reduction occurred via pyruvate:lipoate oxidoreductase. The reaction required catalytic amounts of both coenzyme A and thiamine pyrophosphate. Addition of sodium arsenite inhibited enzyme activity by 90%. Pyruvate:lipoate oxidoreductase was the only system detected in anaerobic, dark-grown R. rubrum cell extracts which operated to produce reduced NAD. The low activity of the enzyme system suggested that it was not quantitatively important in ATP formation.
Effects of low temperature on in vivo and in vitro protein synthesis in Escherichia coli and Pseudomonas fluorescens
Authors: Broeze R J , Solomon C J , Pope D H
Published: January 2020

The effects of temperature on protein synthesis by Escherichia coli, a mesophile, and Pseudomonas fluorescens, a psychotroph, were investigated by using whole-cell and cell extract preparations. After shifts to 5 degrees C, protein was synthesized at a slowly decreasing rate for 1 h by both organisms, after which P. fluorescens synthesized protein at a new rate corresponding to its 5 degrees growth rate, in contrast to E. coli which did not synthesize protein at a measurable rate. In vitro protein-synthesizing systems using MS-2 RNA, endogenous mRNA, and purified polysomes were utilized to investigate initiation of translation at 5 degrees C. In these systems, P. fluorescens cell extracts synthesized protein at linear rates for up to 2 h at 5 degrees C, whereas E. coli cell extracts synthesized protein for only 25 min at 5 degrees C. The rates of polypeptide elongation, as tested by the incorporation of phenylalanine into polyphenylalanine by cell extract protein-synthesizing systems from both organisms, were identical over the range of 25 to 0 degrees C. The polysome profiles of E. coli whole cells shifted from 37 to 5 degrees C showed accumulation of 70S ribosomal particles and ribosomal subunits at the expense of polysomes. Similar experiements done with P. fluorescens resulted in polysome reformation at 5 degrees C. In vitro experiments demonstrated that the 70S ribosomal particles, which accumulated in E. coli at 5 degrees C, were capable of synthesizing protein in vitro in the absence of added mRNA. These in vivo and in vitro results suggest that incubation of E. coli at subminimal temperatures results in a block in initiation of translation causing polysomal runoff and the accumulation of 70S particles, some of which are 70S monosomes.
Cell-Cell Recognition in
Saccharomyces cerevisiae
: Regulation of Mating-Specific Adhesion
Authors: Fehrenbacher George , Perry Karen , Thorner Jeremy
Published: January 2020

Mating-specific adhesion between haploid yeast cells of opposite mating type (
a
and α) was studied by using a quantitative agar plate assay. Washed
a
and α cells that had not previously been exposed to their respective opposite mating type (“naive” cells) adhered relatively weakly. In water, only 5 to 10% of the
a
cells stuck tightly enough to α cells to give rise subsequently to diploid clones on the assay plates. Under optimum conditions (pH 6 to 7, at least 0.1 M Nacl or 0.01 M Mg
2+
), there was about 20% adhesion. Nevertheless, this weak binding defined a mating type-specific interaction because, even under optimum conditions, the homologous interactions (
a
with
a
and α with α) yielded only 3 to 5% cohesion. In contrast to these results, washed cells that had been preincubated in the cell-free culture medium of their opposite mating type (“preconditioned” cells) adhered quite strongly. The degree of adhesion between preconditioned cells (40 to 50%) was essentially unaffected by extremes of ionic strength, pH, and temperature and by the absence of divalent cation. This strong interaction was also mating type specific since cohesion between preconditioned cells of like mating type was only about 5%. The increase in agglutinability was obtained if only the
a
cells were preconditioned and could be induced by highly purified preparations of natural or synthetically prepared α-factor, an oligopeptide pheromone released by the α cells. The appearance of increased adhesiveness was blocked by an inhibitor of RNA synthesis and by an inhibitor of protein synthesis, but not by an inhibitor of polysaccharide synthesis. Adhesion between preconditioned cells could be inhibited by pretreatment with functionally univalent succinylated concanavalin A or with extracts from preconditioned cells of the opposite mating type. These results confirm in a quantitative manner that the recognition between conjugating cells of
S. cerevisiae
is a developmentally regulated event that is under the control of the mating type locus.
NIC, a conjugative nicotine-nicotinate degradative plasmid in Pseudomonas convexa
Authors: Thacker R , Rørvig O , Kahlon P , Gunsalus I C
Published: January 2020

The plasmid nature of genes specifying degradation of nicotine and nicotinate in Pseudomas convexa strain 1 (Pc1) is indicated by mitomycin curing and conjugational transfer to other strains. The NIC plasmid appears to be compatible with other metabolic plasmids in Pseudomonas putida.
Polyethylene glycol-induced fusion of heat-inactivated and living protoplasts of Bacillus megaterium
Authors: Fodor K , Demiri E , Alföldi L
Published: January 2020

Protoplasts of Bacillus megaterium, incubated at 50 degrees C for 120 min, lost the ability to revert to bacillary form. Such heat-inactivated protoplasts, however, produced recombinants when fused by polyethylene glycol treatment with normal protoplasts. Although this differential inactivation effect is not yet fully reproducible, reciprocal inactivations of the parental protoplasts in genetic crosses have clearly shown that for protoplast fusion (i) either of the parents may serve as the viable recipient for markers coming from the heated parental protoplasts, and (ii) either of the parents may be rendered nonviable and yet, when fused with a viable partner, contribute to formation of a recombinant. Heat inactivation seems to provide a way to counterselect when few markers are available and one of the parents is prototrophic.
Enterobacterial common antigen in mutant strains of Salmonella
Authors: Männel D , Mäkelä P H , Mayer H
Published: January 2020

A good correlation was found to exist between the serologically determined presence of enterobacterial common antigen (ECA) and the amount of the rare sugar constituent D-mannosaminuronic acid. Strains classified by serological techniques as ECA+, ECA-, and ECAtrace were found to possess the expected amounts of mannosaminuronic acid in the ECA-enriched phenol-soluble PL-L fractions. This correlation provides strong evidence on the identity of the mannosaminuronic acid-glucosamine polymer with the ECA as defined by Kunin (J. Exp. Med. 118:565-586, 1963).
Chromosomal map location of the methicillin resistance determinant in Staphylococcus aureus
Authors: Kuhl S A , Pattee P A , Baldwin J N
Published: January 2020

Three-factor genetic crosses performed by transformation have shown that the methicillin resistance determinant of Staphylococcus aureus strain DU4916 (the mec-4916 marker) is linked to a novobiocin resistance (Novr) marker (nov-142) and mutational sites affecting pyrimidine (pyr-141), purine (pur-102), and histidine (hisG15) biosynthesis in S. aureus strain 8325. The linkage group thus defined is pyr-141-hisG15-nov-142-pur-102-mec-4916. Phage 80alpha previously propagated on a novobiocin-resistant, methicillin-sensitive (Mecs) 8325 strain was used to infect 21 novobiocin-sensitive, methicillin-resistant clinical isolates (including strain DU4916). Among the novobiocin-resistant transductants so obtained from each recipient, between 1 and 5% were methicillin sensitive (reflecting cotransduction of Novr and Mecs). These results are consistent with the genetic determinant of methicillin resistance having a single chromosomal locus in most, if not all, strains of S. aureus.
Salmonella typhimurium mutants lacking protease II
Authors: Heiman C , Miller C G
Published: January 2020

Mutants of Salmonella typhimurium lacking protease II, an endoprotease with trypsin-like specificity, have been isolated. These mutants can be identified by using the chromogenic substrate N-methyl-N-p-toluenesulfonyl-L-lysine beta-naphthyl ester to screen colonies growing on agar for the presence of the enzyme. All of the mutations isolated map at locus tlp (typsin-like protease) which is cotransducible (approximately 1%) using phage P1 with tre (trehalose utilization) at approximately 58 min on the Salmonella map. Double mutants lacking both protease I and protease II have been constructed. These strains grew normally. They were able to degrade abnormal proteins and to carry out protein turnover during carbon starvation at the same rate as the wild type.
Occurrence of squalene in methanol-grown bacteria
Authors: Goldberg I , Shechter I
Published: January 2020

The nonpolar lipids of methanol-grown bacteria which utilize one-carbon (C1) compounds via the RMP pathway (Pseudomonas C, Pseudomonas methylotropha, and Methylomonas methanolica) were found to contain squalene in concentrations between 0.1 to 1.16 mg/g of cell (dry weight). Squalene could not be detected in lipid extracts of methanol-grown bacteria which utilize C1 compounds via the serine pathway.
Transconjugant analysis: limitations on the use of sequence-specific endonucleases for plasmid identification
Authors: Causey S C , Brown L R
Published: January 2020

We used the sequence-specific endonucleases EcoRI, SmaI, BamHI, HsuI, and HaeIII as identification tools in following the conjugal transfer of the well-studied R plasmids Sa, R388, RP4, and R6K. Transfers were both intergeneric and intrageneric. Plasmid fingerprints were generated from both single- and combination-enzyme digests. The Sa transconjugants yielded plasmids showing consistent fingerprints for each of the respective endonucleases used, whereas the three other R-plasmid transconjugants showed fingerprint changes.
Temperature-dependent variation in the extent of methylation of ribosomal proteins L7 and L12 in Escherichia coli
Authors: Chang F N
Published: January 2020

The amount of epsilon-N-monomethyllysine in ribosomal proteins L7 and L12 in Escherichia coli is dependent upon the cell growth temperature. At 37 degrees C or above, very small amounts were detected. Dramatic increase in the content of epsilon-N-monomethyllysine in these proteins was observed when the growth temperature was lowered.