bluuebunny/crossref_metadata_2025 huggingface dataset:
Respiratory Activities Associated with Mesosomal Vesicles and Protoplast Membranes of
Staphylococcus aureus
Authors: Theodore T. S. , Weinbach E. C.
Published: January 2020

Analysis of oxidase and dehydrogenase activities, cytochrome content of mesosomal vesicles, and protoplast membranes showed that the respiratory chain in
Staphylococcus aureus
is associated predominantly with the protoplast membranes.
Studies with Lectins on the Surface Carbohydrate Structures of Mycoplasma Membranes
Authors: Schiefer Hans-Gerd , Gerhardt Ursula , Brunner Helmut , Krüpe Martin
Published: January 2020

The surface carbohydrate structures on the cell membranes of various mycoplasma species have been investigated by using lectins, which are sugar-specific proteins. Carbohydrate structures presumably bound to glycolipids, with both galactose and glucose units, were found to be exposed on the surface of
Mycoplasma pneumoniae
and its temperature-sensitive mutants,
M. mycoides
var.
mycoides
and
capri, M. pulmonis, M. gallinarum
, and
M. gallisepticum
. Lipid-bound glucose was found on
M. neurolyticum
. The possible relationship of the lipid-bound surface carbohydrate groups to the known serological cross-reactions and lipid compositions of the various mycoplasma species is discussed. Intact
Acholeplasma laidlawii
and
M. fermentans
have no lectin-binding sites exposed on their surfaces; galactose groups were discovered only after Pronase digestion of the organisms, suggesting that their glycolipids are hidden under a protein layer. Neither intact nor Pronase-digested
M. hominis
reacted with the lectins; this is fully consistent with the lipid composition of this organism, which contains glycolipids. The lectins from
Vicia cracca
and
Phaseolus vulgaris
, which react with
N
-acetyl-galactosamine groups, agglutinated
M. gallinarum, M. gallisepticum, M. mycoides
var.
capri
, and
M. pulmonis
. The agglutinability was lost after Pronase treatment, indicating that the corresponding carbohydrates are presumably protein bound. They may be correlated with the extracellular structures observed by electron microscopy of both sectioned and negatively stained mycoplasma species.
Effect of Glucose on the Fruiting Body Formation and Adenosine 3′,5′-Cyclic Monophosphate Levels in
Coprinus macrorhizus
Authors: Uno Isao , Ishikawa Tatsuo
Published: January 2020

The formation of fruiting bodies in the monokaryotic

fis
c

strain and a dikaryon of
Coprinus macrorhizus
was inhibited by growth in high-glucose media. In high-glucose media the characteristic burst of adenosine 3′,5′-cyclic monophosphate (cAMP) accumulation during fruiting-body formation was absent. Enzymatic activity assays revealed that mycelia grown in high-glucose media contained relatively lower amounts of adenylate cyclase and cAMP-phosphodiesterase than mycelia grown in low-glucose media. The synthesis of inducible
d
-serine deaminase and tryptophanase was repressed in high-glucose media. A mutant (
gluR
) in which the glucose repression of fruiting-body formation is affected was isolated by selection in high-glucose media. The mutation caused the cAMP levels to be no longer affected by glucose and affected ability to synthesize the inducible
d
-serine deaminase and tryptophanase. The
gluR
mutant was partially dominant in dikaryons. It is suggested that cAMP may play important roles in inducing fruiting bodies and in controlling inducible enzyme synthesis in
C. macrorhizus
.
Comparison of the Deoxyribonucleic Acid Molecular Weights and Homologies of Plasmids Conferring Linked Resistance to Streptomycin and Sulfonamides
Authors: Barth Peter T. , Grinter Nigel J.
Published: January 2020

Bacterial strains showing linked resistance to streptomycin (Sm) and sulfonamides (Su) were chosen representing a wide taxonomic and geographical range. Their SmSu resistances were transferred to
Escherichia coli
K-12 and then plasmid deoxyribonucleic acid (DNA) was isolated by ethidium bromide CsCl centrifugation. The plasmid DNA was examined by electron microscopy and analyzed by sedimentation through 5 to 20% neutral sucrose gradients. Plasmid DNA from strains having transmissible SmSu resistance consisted of two or three molecular species, one of which had a molecular mass of about 5.7 Mdal (10
6
daltons), the others varying between 20 to 60 Mdal. By using transformation or F′ mobilization, we isolated the SmSu-resistance determinant from any fellow resident plasmids in each strain and again isolated the plasmid DNA. Cosedimentation of each of these with a differently labeled reference plasmid DNA (R300B) showed 9 out of 12 of the plasmids to have a molecular mass not significantly different from the reference (5.7 Mdal); two others were 6.3 and 9.2 Mdal, but PB165 consisted of three plasmids of 7.4, 14.7, and 21.4 Mdal. Three separate isolations of the SmSu determinant from PB165 gave the same three plasmids, which we conclude may be monomer, dimer, and trimer, respectively. DNA-DNA hybridizations at 75 C demonstrated 80 to 93% homology between reference R300B DNA and each isolated SmSu plasmid DNA, except for the 9.2-Mdal plasmid which had 45% homology and PB165 which had 35%. All the SmSu plasmids were present as multiple copies (about 10) per chromosome. The conjugative plasmid of R300 (present as 1.3 copies per chromosome) has been shown to have negligible effect on the number of copies of its accompanying SmSu plasmid R300B. We conclude that the SmSu plasmids are closely related and probably have a common evolutionary origin.
Cold-Induced Increase of Glycerol Kinase Activity in
Neurospora crassa
: Rapid Inactivation of the Enzyme In Vivo
Authors: North M. J.
Published: January 2020

The glycerol kinase activity induced by incubation of
Neurospora crassa
at low temperatures was rapidly lost when cultures were returned to 26 C. After a short lag, the activity disappeared irreversibly with a half-life of approximately 15 min. The loss of activity was not due to a change in the level of an inhibitor or activator. Glycerol reduced the activity loss but did not completely prevent it, which was an effect that was dependent on protein synthesis. The cold-induced activity was also always lost on addition of cycloheximide at all temperatures tested (0 to 26 C), which indicated continuous inactivation, although cycloheximide did not affect the actual rate of activity loss at 26 C. The basal glycerol kinase activity was not sensitive to cycloheximide. The mechanism responsible for inactivation was destroyed by sonic oscillation. The process is not thought to play a role in the cold-induced increase in activity. Glycerol kinase activity induced at 26 C by glycerol was also lost on addition of cycloheximide and after addition of sucrose.
TherepMutation III. Altered Structure of the ReplicatingEscherichia coliChromosome
Authors: Lane H. E. David , Denhardt David T.
Published: January 2020

Therepgene function ofEscherichia coliis essential for the replication of P2 and φX174 double-stranded deoxyribonucleic acid (DNA). Compared with isogenicrep+strains,repmutants show the following characteristics: larger cell size, more DNA per cell, and a slightly lower DNA/mass ratio. The replicatingrepchromosomes show a steeper gradient of marker frequencies and contain more replicating forks per chromosome. The nucleoid body ofrepmutants sediments faster and contains more DNA. We deduce that therepfunction is required for the “normal” replication of theE. colichromosome and that in its absence theE. colichromosome replicates in an altered manner, perhaps involving slower-moving replicating forks.
Stimulation of Transport into
Escherichia coli
Membrane Vesicles by Internally Generated Reduced Nicotinamide Adenine Dinucleotide
Authors: Futai Masamitsu
Published: January 2020

Membrane vesicles containing internal alcohol dehydrogenase and nicotinamide adenine dinucleotide (NAD) were prepared from
Escherichia coli
ML308-225. Upon the addition of ethanol, these vesicles respired and transported several amino acids. Transport rates driven by internally generated reduced NAD (NADH) were about 60 to 80% of that stimulated by
d
-lactate. This transport was inhibited by cyanide and anaerobiosis. Ferricyanide, a nonpermeable electron acceptor, inhibited transport stimulated by external NADH, but not that stimulated by internally generated NADH.
Purification and Characterization of the Two 6-Phosphogluconate Dehydrogenase Species from
Pseudomonas multivorans
Authors: Lee Young Nam , Lessie T. G.
Published: January 2020

The two species of 6-phosphogluconate dehydrogenase (EC 1.1.1.43) from
Pseudomonas multivorans
were resolved from extracts of gluconate-grown bacteria and purified to homogeneity. Each enzyme comprised between 0.1 and 0.2% of the total cellular protein. Separation of the two enzymes, one which is specific for nicotinamide adenine dinucleotide phosphate and the other which is active with nicotinamide adenine dinucleotide or nicotinamide adenine dinucleotide phosphate was facilitated by the marked difference in their respective isoelectric points, which were at pH 5.0 and 6.9. Comparison of the subunit compositions of the two enzymes indicated that they do not share common peptide chains. The enzyme active with nicotinamide adenine dinucleotide was composed of two subunits of about 40,000 molecular weight, and the nicotinamide adenine dinucleotide phosphate-specific enzyme was composed of two subunits of about 60,000 molecular weight. Immunological studies indicated that the two enzymes do not share common antigenic determinants. Reduced nicotinamide adenine dinucleotide phosphate strongly inhibited the 6-phosphogluconate dehydrogenase active with nicotinamide adenine dinucleotide by decreasing its affinity for 6-phosphogluconate. Guanosine-5'-triphosphate had a similar influence on the nicotinamide adenine dinucleotide phosphate-specific 6-phosphogluconate dehydrogenase. These results in conjunction with other data indicating that reduced nicotinamide adenine dinucleotide phosphate stimulates the conversion of 6-phosphogluconate to pyruvate by crude bacterial extracts suggest that in
P. multivorans
, the relative distribution of 6-phosphogluconate into the pentose phosphate and Entner-Doudoroff pathways might be determined by the intracellular concentrations of reduced nicotinamide adenine dinucleotide phosphate and purine nucleotides.
Construction of φ80 dhisCarryingSalmonella typhimuriumHistidine Operon Mutations
Authors: Smith Gerald R. , Tong Benton
Published: January 2020

Starting with a previously isolated F′hisepisome and φ80 dhis immλcI857transducing phage, we have constructed recombinant elements bearing previously isolatedhismutations fromSalmonella typhimurium. These phages were constructed as sources of deoxyribonucleic acid for in vitro biochemical experiments on gene regulation. The manipulation of genes betweenS. typhimuriumandEscherichia colidescribed here may be useful in studying otherS. typhimuriumoperons.
Involvement of Threonine Dehydratase in Biosynthesis of the α-Ketobutyrate Prosthetic Group of Urocanase
Authors: Daumy G. O. , Cohn M. S. , Phillips A. T.
Published: January 2020

Seventeen mutants of
Pseudomonas putida
that were unable to grow on threonine as nitrogen source owing to a lack of threonine dehydratase were isolated, and all were found to be unable to synthesize active urocanase. Spontaneous revertants selected for urocanase production concomitantly regained threonine dehydratase. Mutants that were unable to utilize urocanate as carbon source were also isolated, and these were defective in urocanase formation but were normal in threonine dehydratase levels. Since α-ketobutyrate is the prosthetic group for urocanase, these results are consistent with the proposal that threonine dehydratase is necessary for urocanase prosthetic group biosynthesis. However, the lack of urocanase activity in threonine dehydratase-negative mutants was shown not to be the result of reduced levels of endogenous free α-ketobutyrate, nor to the participation of threonine dehydratase in the initiation of urocanase biosynthesis through the conversion of threonyl-tRNA
Thr
to α-ketobutyryl-tRNA
Thr
. Other alternatives for the participation of threonine dehydratase in urocanase biosynthesis are discussed.
Ribonucleic Acid Regulation in Amino Acid-Limited Cultures of
Escherichia coli
Grown in a Chemostat
Authors: Atherly Alan G.
Published: January 2020

The regulation of ribonucleic acid (RNA) synthesis was examined in cultures of bacteria whose growth was limited in the chemostat by the supply of a required amino acid. Strains possessing the relaxed (
relA
) mutation accumulated excess RNA (relative to protein) at low growth rates when growth was limited by arginine, histidine, or cysteine but not when limited by methionine. In contrast, stringent (
relA
+
) strains maintained a constant RNA/protein ratio with decreasing growth rate regardless of the amino acid used to limit growth. The presence of excess RNA in relaxed strains was accompanied by an absence of increase in RNA production upon addition of chloramphenicol, a lag upon shift-up in growth by addition of excess of the limiting amino acid, and a decreased rate of production of β-galactosidase upon induction. Analysis of the RNA accumulated in relaxed strains indicated it was present as transfer RNA as well as 50
S
and 30
S
ribosomal subunits. Microscope examination of the relaxed strains during histidine-, arginine-, or cysteine-limited growth in the chemostat showed them to be 10 to 20 times longer in size than the stringent strains. Also, cell density was reduced to one-tenth when the increased size was observed. An analysis of the amount of ppGpp present in all slow-growing amino acid-limited cultures (relaxed and stringent) demonstrated that only basal levels of ppGpp were made. These data are consistent with the hypothesis that when growth is limited in the chemostat by an initiation event in protein synthesis, i.e., limited methionine, RNA regulation occurs in relaxed as well as stringent strains. Also, when other amino acids are limiting in concentration during translation, errors occur in relaxed strains, resulting in misread proteins.
Characterization of the lipids of mesosomal vesicles and plasma membranes from Staphylococcus aureus
Authors: Beining P R , Huff E , Prescott B , Theodore T S
Published: January 2020

Mesosomal vesicles and plasma membranes were isolated from Staphylococcus aureus ATCC 6538P by protoplasting and differential centrifugation. The lipids of each of the two membrane fractions were extracted with pyridine-acetic acid-N-butanol, and the nonlipid contaminants were removed by Sephadex treatment. The lipids were then separated by passage through diethylaminoethyl-cellulose columns and characterized by thin-layer chromatographic, chemical, and spectral analyses. The lipids were separated into four discrete diethylaminoethyl fractions: (i) vitamin K2, carotenoids, C55 isoprenoid alcohol, and monoglucosyl diglyceride; (ii) cardiolipin, carotenoids, phosphatidyl glycerol, diglucosyl diglyceride, and an unidentified ninhydrin-positive component; (iii) cardiolipid and phosphatidyl glyderol; (iv) cardiolipin, phosphatidyl glycerol, and phosphatidyl glucose. Qualitatively, no difference in lipid composition between mesosomal vesicles and plasma membranes was found. However, based on equal dry weights of membrane materials, a relative quantitative difference in the amount of specific lipids in mesosomal vesicles and plasma membranes was observed. There are 4 times more monoglucosyl diglyceride, 2.6 times more diglucosyl diglyceride, 3.8 times more phosphatidyl glucose, 2 times more carotenoids, and 2 times more vitamin K2 found in mesosomal vesicles than in plasma membranes. The concentration of cardiolipin and phosphatidyl glycerol is 3.6 and 6 times greater, respectively, in mesosomal vesicles.
Transport of D- and L-tryptophan in Bacillus megaterium by an inducible permease
Authors: Bouknight R R , Sadoff H L
Published: January 2020

Tryptophan-grown cells of Bacillus megaterium ATCC 19213 contain a permease system that transports both D- and L-tryptophan and is inhibited by sodium azide. Arginine-grown cells contain little tryptophan permease activity, suggesting that the system is inducible. Arginine represses the tryptophan permease as well as the transport system for leucine and phenylalanine. Kynurenine was a more effective inducer of the tryptophan transport system than either D- or L-tryptophan.
Blocking of bacteriophages phi W and phi 5 with lipopolysaccharides from Escherichia coli K-12 mutants
Authors: Boman H G , Monner D A
Published: January 2020

In the preceding paper we presented a formula for the composition of lipopolysaccharides (LPS) from Escherichia coli K-12. This formula contains four regions defined from analyses of LPS from four key strains, the parent and mutants which had lost one, two, or three regions of their carbohydrates. Support for the formula was derived from the susceptibility of the key mutants to several bacteriophages. One of these, phage phi W, was found specific for strains which had lost region 4. In this paper we described inactivation in vitro of phage phi W and its host-range mutant phi 5, using LPS devoid of regions 2 to 4. The blocking of phi W was found to require about 0.15 M concentrations of monovalent cations and to be inhibited by low concentrations of calcium and magnesium ions. One particle of phage phi W required 2 times 10-16 g of LPS devoid of region 4 for stoichiometric inactivation. Phage phi 5 was blocked by both heptose-less LPS (devoid of regions 2 to 4) and glucose-less LPS (devoid of regions 3 to 4) but was unaffected by LPS devoid of region 4. LPS from a heptose-less mutant of Salmonella minnesota showed the same inactivation ability as did LPS from heptose-less strains of E. coli K-12. Lipid A was prepared from LPS containing all four regions. Such lipid A was found to inactivate phi 5, whereas both the polysaccharide moiety as well as the intact LPS were without effect. It is suggested that lipid A is part of the receptor site for phage phi 5.
Urea transport in Saccharomyces cerevisiae
Authors: Cooper T G , Sumrada R
Published: January 2020

Urea transport in Saccharomyces cerevisiae occurs by two pathways. The first mode of uptake is via an active transport system which: (i) has an apparent Km value of 14 muM, (ii) is absolutely dependent upon energy metabolism, (iii) requires pre-growth of the cultures in the presence of oxaluric acid, gratuitous inducer of the allantoin degradative enzymes, and (iv) is sensitive to nitrogen repression. The second mode of uptake which occurs at external urea concentrations in excess of 0.5 mM is via either passive or facilitated diffusion.
Production of racemic lactic acid in Pediococcus cerevisiae cultures by two lactate dehydrogenases
Authors: Gordon G L , Doelle H W
Published: January 2020

Nicotinamide adenine dinucleotide (NAD)-dependent d(minus)-and l(plus)-lactate dehydrogenases have been partially purified 89- and 70-fold simultaneously from cell-free extracts of Pediococcus cerevisiae. Native molecular weights, as estimated from molecular sieve chromatography and electrophoresis in nondenaturing polyacrylamide gels, are 71,000 to 73,000 for d(minus)-lactate dehydrogenase and 136,000 to 139,000 for l(plus)-lactate dehydrogenase. Electrophoresis in sodium dodecyl sulfate-containing gels reveals subunits with approximate molecular weights of 37,000 to 39,000 for both enzymes. By lowering the pyruvate concentration from 5.0 to 0.5 mM, the pH optimum for pyruvate reduction by d(minus)-lactate dehydrogenase decreases from pH 8.0 to 3.6. However, l(plus)-lactate dehydrogenase displays an optimum for pyruvate reduction between pH 4.5 and 6.0 regardless of the pyruvate concentration. The enzymes obey Michaelis-Menten kinetics for both pyruvate and reduced NAD at pH 5.4 and 7.4, with increased affinity for both substrates at the acid pH. alpha-Ketobutyrate can be used as a reducible substrate, whereas oxamate has no inhibitory effect on lactate oxidation by either enzyme. Adenosine triphosphate causes inhibition of both enzymes by competition with reduced NAD. Adenosine diphosphate is also inhibitory under the same conditions, whereas NAD acts as a product inhibitor. These results are discussed with relation to the lactate isomer production during the growth cycle of P. cerevisiae.
Changes in composition, biosynthesis, and physical state of membrane lipids occurring upon aging of Mycoplasma hominis cultures
Authors: Rottem S , Greenberg A S
Published: January 2020

During the progression of Mycoplasma hominis cultures from the early logarithmic phase to the stationary phase of growth, the total phospholipid content of the cell membranes decreased. Measurement of the amount of the various phospholipids during the growth cycle showed that a decrease in the phosphatidylglycerol (PG) content, accompanied by an increase in the phosphatidic acid content, occurred upon aging of the culture. Pulse labeling experiments revealed that the PG, once formed, is relatively stable, undergoing no detectable turnover in growing cultures of M. hominis. No changes in the fatty acid composition of the membrane phospholipids were observed on aging of the culture, with palmitic acid predominating throughout the growth cycle. The preferential incorporation of palmitate into the phospholipid fraction is apparently caused by the higher activity of the membrane-bound acyl-coenzyme A (CoA):alpha-glycerophosphate transacylase with palmityl-CoA rather than with oleyl-CoA as substrate. The activity of the soluble acyl-CoA synthetase was the same whether palmitate or oleate served as substate. M. hominis membrane preparations contained a PG-synthetase system catalyzing the incorporation of L-alpha-glycerol-3-phosphate into PG. The activity of the PG synthetase system was markedly dependent on the age of the culture, being highest in cells from the early exponential phase of growth while declining sharply thereafter, and thus probably responsible, in part, for the decrease in PG content upon aging of the cells. Electron paramagnetic resonance spectra of a spin-labeled fatty acid incorporated in M. hominis membranes revealed a marked decrease in the freedom of motion of the spin label on aging of the culture. It is proposed that this decrease is due primarily to the decrease in the lipid-to-protein ratio of the membranes and has a marked effect on the activity of membrane-associated enzymes and transport systems.
Metabolism of pyrimidine deoxyribonucleosides in Neurospora crassa
Authors: Shaffer P M , Hsu C A , Abbott M T
Published: January 2020

The experiments in this report involve the following series of reactions which were previously demonstrated with purified enzyme preparations from Neurospora crassa: thymidine a yields thymine ribonucleoside b yields thymine c yields 5-hydroxymethyluracil d yields 5-formyluracil e yields uracil-5-carboxylic acid f yields uracil. The evidence for some of the reactions occurring in vivo has been incomplete and for others totally lacking. In this paper intact cells of Neurospora are shown to be capable of converting the substrates of each of the reactions to the corresponding products. Studies are described which were carried out in vivo and in vitro with the pyrimidineless strains pyr-4,uc-1,uc-2 and pyr-4,uc-1,uc-3, developed by Williams and Mitchell. The results reported in the present paper indicate that (reaction a) and the uc-3 mutation affects thymine 7-hydroxylase (reactions c,d, and e). Evidence is presented for the 2'-hydroxylase reaction being the major, if not only, way by which Neurospora can initiate the conversion of thymidine to the pyrimidines of nucleic acids and for the 2'-hydroxylation of thymidine and deoxyuridine being catalyzed by the same enzyme. Deoxycytidine was shown not to be hydroxylated in intact cells but instead deaminated to deoxyuridine, which in turn was converted to uridine. Further studies with the uc-3-carrying strain showed that an enzyme other than thymine 7-hydroxylase can also convert 5-formyluracil to uracil-5-carboxylic acid.
Solubilization of Escherichia coli nitrate reductase by a membrane-bound protease
Authors: MacGregor C H
Published: January 2020

Nitrate reductase extracted from the membrane of Escherichia coli by alkaline heat treatment was purified to homogeneity and used to prepare specific antibody. Nitrate reductase, precipitated by this antibody from Triton extracts of the membrane, contained a third subunit not present in the purified enzyme used to prepare the antibody. Nitrate reductase precipitated by antibody from alkaline heat extracts was composed of peptide fragments of various sizes. These fragments were produced by a membrane-bound protease which was activated by alkaline pH and heat. It is the action of this protease that releases the enzyme from the membrane, as shown by the observations that protease inhibitors decreased the amount of solubilization of the enzyme, and the enzyme remaining in the membrane after heating showed much less proteolytic cleavage than that which was released.
Nuclear and cell division in Bacillus subtilis: dormant nucleoids in stationary-phase cells and their activation
Authors: Van Iterson W , Michels P A , Vyth-Dreese F , Aten J A
Published: January 2020

The morphology of nucleoids and mesosomes of Bacillus subtilis in stationary-and lag-phase cultures was studied by making three-dimensional cell reconstructions in plastic of electron micrographs of serial sections. In cells from stationary cultures, the dormant nucleoids are frequently, but not always, spherical and the mesosomes are small and compact. It is suggested that the spherical nucleoids represent the resting stage in which replication and segregation have been completed. In cells from lag-phase cultures, the compact mesosomes develop into an elaborate system of tubes and wider sacs which become wrapped around the elongating nucleoids and invade the nucleoplasm in preparation for division.
Regulation of the L-lactase dehydrogenase from Lactobacillus casei by fructose-1,6-diphosphate and metal ions
Authors: Holland R , Pritchard G G
Published: January 2020

The lactate dehydrogenase of Lactobacillus casei, like that of streptococci, requires fructose-1,6-diphosphate (FDP) for activity. The L. casei enzyme has a much more acidic pH optimum (pH 5.5) than the streptococcal lactate dehydrogenases. This is apparently due to a marked decrease in the affinity of the enzyme for the activator with increasing pH above 5.5; the concentration of FDP required for half-maximal velocity increase nearly 1,000-fold from 0.002 mM at pH 5.5 to 1.65 mM at 6.6. Manganous ions increase the pH range of activity particularly on the alkaline side of the optimum by increasing the affinity for FDP. This pH dependent metal ion activation is not specific for Mn2+. Other divalent metals, Co2+, Cu2+, Cd2+, Ni2+, Fe2+, Fe2+, and Zn2+ but not Mg2+, will effectively substitute for Mn2+, but the pH dependence of the activation differs with the metal ion used. The enzyme is inhibited by a number of commonly used buffering ions, particularly phosphate, citrate, and tris (hydroxymethyl) aminomethane-maleate buffers, even at low buffer concentrations (0.02 M). These buffers inhibit by affecting the binding of FDP.
Relation between reduced nicotinamide adenine dinucleotide oxidation and amino acid transport in membrane vesicles from Bacillus subtilis
Authors: Bisschop A , de Jong L , Lima Costa M E , Konings W N
Published: January 2020

The rate of reduced nicotinamide adenine dinucleotide (NADH) oxidation by membrane vesicles from Bacillus subtilis W23 increases three- to fourfold during logarithmic growth, reaching maximal levels in early stationary phase. Initial rates of L-proline, L-alanine, and L-glutamate transport energized by NADH closely parallel the increase in NADH oxidation. In vesicles prepared at different stages of growth, a constant number of NADH molecules varying from 150 to 260 have to be oxidized to transport one molecule of amino acid. Membrane vesicles from B. subtilis aroD (strain RB163), a mutant defective in menaquinone synthesis, do not transport amino acids in the presence of NADH. Ascorbate plus phenazine methosulfate, however, energizes amino acid transport equally well as in vesicles of B. subtilis W23. NADH oxidation and NADH-driven amino acid transport can be restored instantaneously by the addition of menadione (vitamin K3).
Self-transferable plasmids determining the hemolysin and bacteriocin of Streptococcus faecalis var. zymogenes
Authors: Jacob A E , Douglas G J , Hobbs S J
Published: January 2020

Strains of Streptococcus faecalis var. zymogenes, designated JH1 and JH3, produced a hemolysin and a bacteriocin. Hemolytic activity was lost from a low percentage of cells grown in broth at either 37 or 45 C. All nonhemolytic (Hly-) variants had lost bacteriocin activity (Ben-), and those from strain JH3 had also lost resistance to the bacteriocin (Bnr-). The majority of Hly-, Ben- variants from JH1 retained bacteriocin resistance (Bnrplus). Strains JH1 and JH3 contained a plasmid deoxyribonucleic acid species of molecular weight 38 times 10-6 (plasmids pJH2 and pJH3, respectively), and strain JH1 also contained a 50 times 10-6 molecular weight plasmid (pJH1) which has previously been shown to carry the genes determining resistance to the antibiotics kanamycin, neomycin, streptomycin, erythromycin, and tetracycline. Hly-, Bcn-, Bnr- variants of strain JH3 had completely lost plasmid pJH3. Hly-, Bcn-, Bnr- variants of strain JH1 had completely lost plasmid pJH2 and retained plasmid pJH1, but Hly-, Bcn-, Bnrplus variants had retained both plasmids pJH2 and pJH1. The Hlyplus, Bcnplus, Bnrplus traits from both parental strains were transferable to nonhemolytic S. faecalis strains during mixed incubation in broth at 37 C, and hemolytic recipient strains were found to have received plasmid pJH2 from strain JH1 and pJH3 from JH3. We conclude that the Hlyplus, Bnrplus traits are borne on plasmid pJH2 in strain JH1 and pJH3 in strain JH3 and that, in Hly-, Bcn-, Bnrplus variants of strain JH1, plasmic pJH2 has suffered a mutation affecting hemolysin and bacteriocin expression. We infer that the plasmids transfer by conjugation. Beta-hemolytic activity is the only property distinguishing the zymogenes variety from S. faecalis. Since we have shown that this activity is plasmid borne in strains JH1 and JH3, we endorse the view that the varietal status of zymogenes should be dropped.
Purification and properties of a serine protease from Pseudomonas matophilia
Authors: Boethling R S
Published: January 2020

The extracellular protease of Pseudomonas maltophilia was partially purified by ammonium sulfate precipitation and chromatography on Sephadex G-75 and Bio-rex 70. Gel electrophoresis revealed minor impurities. The enzyme exhibited the following properties: (i) molecular weight, 35,000; (ii) A see article; 10.8; (iii) isoelectric point, 9.3; (iv) pH optimum, 10.0; (v)s20, w equal 3.47. The enzyme was rapidly inactivated by ethylenediaminetetracetate, but activity could be partially restored with divalent cations. Of those tested, Ca2+, Sr2+, Ba2+, Co2+, Cu2+, Mg2+, and Zn2+ were all effective. Both phenylmethylsulfonylfluoride and diisopropylfluorophosphate were powerful inhibitors of protease activity, but L-1-tosylamide-2-phenylethylchloromethyl ketone, iodoacetic acid, and iodoacetamide were without effect. The enzyme hydrolyzed the esters N-acetyl-L-tyrosine ethyl ester and alpha-N-benzoyl-L-arginine ethyl ester (BAEE) with Km values of 10.4 and 3.4 mM, respectively. The hydrolysis of BAEE was also inhibited by phenylarsonic acids. The kinetics of inhibition by m-nitrophenylarsonate were of the mixed type, and the K1 was 1.8 mM. The data followed a theoretical curve for a 1:1 enzyme-inhibitor complex with a dissociation constant of 1.8 mM. Inhibition by m-nitrophenylarsonate was pH dependent and followed a theoretical curve for the titration of a protonated group with a pKa of 7.0.
Regulation of the tricarboxylic acid cycle in gram-positive, facultatively anaerobic bacilli
Authors: Tanaka N , Hanson R S
Published: January 2020

The facultative anaerobes Bacillus polymyxa Hino G, B. polymyxa Hino J, and B.macerans were observed to have imcomplete tricarboxylic acid cycles. They were devoid of malate dehydrogenase and all had very low levels of alpha-ketoglutarate dehydrogenase. B. polymyxa Hino J was devoid of alpha-ketoglutarate dehydrogenase when grown aerobically and anerobically. Citrate synthase from B. polymyxa was inhibited by adenosine triphosphate but not reduced nicotinamide adenine dinucleotide and resembled enzymes from other gram-positive bacteria in this respect. Like the citrate synthases from gram-negative, facultative anaerobes and chemolithotrophs, the enzyme from B. polymyxa was inhibited by alpha-ketoglutarate. Inhibition by adenosine triphosphate was shown to be competitive with acetyl-coenzyme A and alpha-ketoglutarate inhibition was competitive with oxaloacetate.
Regulation of the dicarboxylic acid part of the citric acid cycle in Bacillus subtilis
Authors: Ohné M
Published: January 2020

The regulation of alpha-ketogluterate dehydrogenase, succinate dehydrogenase, fumarase, malate dehydrogenase, and malic enzyme has been studied in Bacillus subitilis. The levels of these enzymes increase rapidly during late exponential phase in a complex medium and are maximal 1 to 2 h after the onset of sporulation. Regulation of enzyme synthesis has been studied in the wild type and different citric acid cycle mutants by adding various metabolites to the growth medium. Alpha-ketoglutarate dehydrogenase is induced by glutamate or alpha-ketoglutarate; succinate dehydrogenase is repressed by malate; and fumarase and malic enzyme are induced by fumarate and malate, respectively. The addition of glucose leads to repression of the citric acid cycle enzymes whereas the level of malic enzyme is unaffected. Studies on the control of enzyme activities in vitro have shown that alpha-ketoglutarate dehydrogenase and succinate dehydrogenase are inhibited by oxalacetate. Enzyme activities are also influenced by the energy level, expressed as the energy charge of the adenylate pool. Isocitrate dehydrogenase, alpha-ketoglutarate dehydrogenase, succinate dehydrogenase, and malic enzyme are inhibited at high energy charge values, whereas malate dehydrogenase is inhibited at low energy charge. A survey of the regulation of the citric acid cycle in B.subtilis, based on the present work and previously reported results, is presented and discussed.
Amino acid transport in membrane vesicles of obligately anaerobic Veillonella alcalescens
Authors: Konings W N , Boonstra J , De Vries W
Published: January 2020

Membrane vesicles of Veillonella alcalescens, grown in the presence of L-lactate and KNO-3, actively transport amino acids under anaerobic conditions in the presence of several electron donors and the electron acceptor nitrate. The highest initial rates of uptake are obtained with L-lactate, followed by reduced nicotinamide adenine dinucleotide, glycerol-1-phosphate, formate, and L-malate.. The membrane vesicles contain the dehydrogenases for these electron donors, and these enzymes are coupled with nitrate reductase. In membrane vesicles from cells, grown in the presence of nitrate, the dehydrogenases are not coupled with fumarate reducatase, and anaerobic transport of amino acids does not occur with fumarate as electron acceptor. Under aerobic conditions none of the physiological electron donors can energize transport. However, a high rate of uptake is observed with the electron donor system ascorbate-phenazine metho-sulfate. This electron donor system also effectively energizes transport under anaerobicconditions in the presence of the electron acceptor nitrate.
Origin of the TEM-beta-lactamase gene found on plasmids
Authors: Heffron F , Sublett R , Hedges R W , Jacob A , Falkow S
Published: January 2020

A sequence of deoxyribonucleic acid of 2.7 times 10-6 to 3.3 times 10-6 daltons which includes the TEM beta-lactamase gene is present on the small plasmid RSF 1030 (R-Amp). This same sequence is present on plasmid derivatives that have received a translocation of deoxyribonucleic acid specifying the TEM beta-lactamase and is also present on naturally occurring plasmids of the F1, F11, N, X, O, I, C, and W incompatibility groups that do not specify ampicillin resistance or specify O-type beta-lactamases.
Indirect selection for plasmid mutants: isolation of ColVBtrp mutants defective in self-maintenance in Escherichia coli
Authors: Koyama A H , Wada C , Nagata T , Yura T
Published: January 2020

An efficient method for isolation of a large number of plasmid mutants is described. It is based on the fact that N-methyl-N'-nitro-N-nitrosoguanidine induces a number of closely linked mutations within a short segment of the bacterial chromosome. Thus, selection for reversions of an auxotrophic marker located on the ColVBtrp plasmid yielded a large fraction (more than 50 percent) of mutants defective in some plasmid functions, including its own maintenance in the host bacteria. The results of preliminary characterization of strains carrying these mutated plasmids are presented.
Two distinct isocitrate lyases from a pseudomonas species
Authors: Bellion E , Woodson J
Published: January 2020

The isocitrate lyases of acetate- and methylamine-grown Pseudomonas MA (Shaw strain) were studied. They were shown to be different by a variety of physical criteria including chromatographic elution patterns, heat inactivation kinetics, pH variation of Km values, and migration on polyacrylamide gels. The implications and significance of the existence of two enzymes in relation to the role of isocitrate lyase in methylamine utilization is discussed.
Morphological change in the early stages of the mating process of Rhodosporidium toruloides
Authors: Abe K , Kusaka I , Fukui S
Published: January 2020

The events which occur in the early stages of the mating process of the yeast Rhodosporidium toruloides between strains M-919 (mating type A) and M-1057 (mating type a) were investigated. In preliminary experiments we determined the frequency of mating by two newly designed methods: the liquid culture method and the membrane-filter microculture method. The mating frequencies of strains M-919 and M-1057 were 89% in the liquid culture method and 62% in the membrane-filter microculture method. The early stages in the mating process included the following events: (i) M-919 cells produce constitutively the extracellular inducing substance (A factor), (ii) M-1057 cells receive A factor, and in response to it they form mating tubes and secrete another inducing substance (a factor), (iii) M-919 cells receive a factor, and in response to it they form mating tubes, (iv) mating tubes elongate to the cells or the tubes of mating partner, (v) tips of the growing tubes recognize the opposite mating type cells or their tubes, followed by cell-to-cell fusion.
Regulation of Glutamine Transport in Escherichia coli
Authors: Willis R C , Iwata K K , Furlong C E
Published: January 2020

The formation of the high-affinity (Km equal to 0.2 muM) L-glutamine transport system of Escherichia coli strain 7 (Lin) appears to be subject to the same major control as the glutamine synthetase (EC 6.3.1.2) of this gram-negative organism. Culture of cells under nitrogen-limited conditions provides maximum derepression of both the glutamine synthetase and the glutamine transport system. Nutritional conditions providing a rich supply of ammonium salts or available sources of nitrogen, i.e., conditions which repress the formation of glutamine synthetase, provide three- and 20-fold repression, respectively, of the glutamine transport system. Culture of cells with glutamine supplements of 2 mM does not increase the repression of high-affinity glutamine transport system beyond the level observed in the absence of glutamine. A second kinetically distinct low-affinity component of glutamine. A second kinetically distinct low-affinity component of glutamine uptake is observed in cells cultured with a glutamine-depleted nutrient broth. This second component is associated with the appearance of glutaminase A (EC 3.5.1.2) and asparaginase I (EC 3.5.1.1), a periplasmic enzyme. Parallel changes were observed in the levels of the high-affinity glutamine transport system and the glutamine synthetase when cells were cultured with the carbon sources: glucose, glycerol, or succinate.
Translational Control of Protein Synthesis in Staphylococcus aureus
Authors: Martin S E , Iandolo J J
Published: January 2020

The calculated in vivo polypeptide chain growth rate for Staphylococcus auteus MF-31 grown in nutritionally rich medium assuming all the ribosomes were functional was found to be approximately 16 amino acids/s/ribosome, but decreased to 10.2 amino acids/s/ribosome for cells grown in poor medium. An in vitro analysis revealed that 70S ribosomes isolated from rich medium cells were more active than similar 70S ribosomes derived from cells grown in poor medium. The 30S subunit was found responsible for the increased activity of the rich monosomes, whereas the 50S subunit appeared to be capable of either high or low activity.
chlD gene function in molybdate activation of nitrate reductase
Authors: Sperl G T , DeMoss J A
Published: January 2020

chlD mutants of Escherichia coli lack active nitrate reductase but form normal levels of this enzyme when the medium is supplemented with 10-3 M molybdate. When chlD mutants were grown in unsupplemented medium and then incubated with molybdate in the presence of chloramphenicol, they formed about 5% the normal level of nitrate reductase. Some chlD mutants or the wild type grown in medium supplemented with tungstate accumulated an inactive protein which was electrophoretically identical to active nitrate reductase. Addition of molybdate to those cells in the presence of chloramphenicol resulted in the formation of fully induced levels of nitrate reductase. Two chlD mutants, including a deletion mutant, failed to accumulate the inactive protein and to form active enzyme under the same conditions. Insertion of 99-Mo into the enzyme protein paralleled activation; 185-W could not be demonstrated to be associated with the accumulated inactive protein. The rates of activation of nitrate reductase at varying molybdate concentrations indicated that the chlD gene product facilitates the activation of nitrate reductase at concentrations of molybdate found in normal growth media. At high concentrations, molybdate circumvented this function in chlD mutants and appeared to activate nitrate reductase by a mass action process. We conclude that the chlD gene plays two distinguishable roles in the formation of nitrate reductase in E. coli. It is involved in the accumulation of fully induced levels of the nitrate reductase protein in the cell membrane and it facilitates the insertion of molybdenum to form the active enzyme.
Genetic and physical studies of lambda transducing bacteriophage carrying the beta subunit gene of the Escherichia coli ribonucleic acid polymerase
Authors: Ikeuchi T , Yura T , Yamagishi H
Published: January 2020

The prophage lambdac1857 was inserted into the bfe gene located near rif (the structural gene for the beta subunit of deoxyribonucleic acid [DNA]-dependent ribonucleic acid polymerase) on the Escherichia coli chromosome. Induced lysates (low-frequency transducing lysates) of such a lysogen contained defective lambda phage particles (lambdadrif+) that can specifically transduce the wild-type rif+ gene. Upon transduction into a recipient strain carrying recA, heterogenotes harboring both the wild-type and the mutant rif genes were isolated. Rec+ derivatives of these heterogenotes produce high-frequency transducing lysates that contain lambdadrif+ and normal active phages at a ratio of 1 to 2. The results of marker rescue experiments and of density determination with several transducing phages indicate that most of the late genes are deleted and replaced by a segment of the chromosomal DNA carrying the bfe-rif region. The length of the chromosomal segment seems to vary between approximately 0.5 and 0.6% of the total bacterial DNA among the three independently isolated lambdadrif+ phages. Electron microscopy of heteroduplex DNA consisting of one strand from lambdadrif+-6 and the other from lambdaimm-21 phages directly confirmed that most of the phage DNA of the "left arm" was replaced by the bacterial DNA. The heteroduplex study also demonstrated that the integration of prophage lambda into the bfe region occurred at the normal cross-over point within the phage attachment site.
Inhibition of an early event in the cell division cycle of Escherichia coli by FL1060, an amidinopenicillanic acid
Authors: James R , Haga J Y , Pardee A B
Published: January 2020

Analysis of exponential and synchronous cultures of Escherichia coli B/r after the addition of FL1060 indicates a block point for division by this agent some 15 to 20 min before the end of the preceding cell division cycle, a time corresponding to the beginning of the C period of the cell division cycle. Morphological examination of FL1060-treated synchronous cultures of E. coli /r was consistent with inhibition by FL1060 of a very early event in the cell division cycle. This event appears to be essential for normal cell surface elongation in a rod configuration. Temporary treatment of synchronous cultures of E. coli B/r with FL1060 resulted in division delay, the extent of which was a function of the duration of exposure to FL1060. However, even after relatively long times of FL1060 treatment the delayed divisions were still synchronous. Although FL1060 had no direct effect on deoxyribonucleic acid (DNA) synthesis, the synchronous delayed division occuring after temporary treatment with FL1060 were accompanied by a delay in the attainment of resistance of cell division to inhibitors of DNA, ribonucleic acid, and protein synthesis. These results suggest aht an FL1060-sensitive event initiates at the beginning of the C period of the cell division cycle of E. coli and is responsible for normal cell elongation. This cell elongation pathway procedes independently of DNA synthesis, but there is an interaction between this pathway and termination of a round of DNA replication in which a normal rod configuration is necessary to allow a signal for cell division to be generated upon completion of DNA replication.
Oxidation of C1 Compounds by Particulate fractions from Methylococcus capsulatus: distribution and properties of methane-dependent reduced nicotinamide adenine dinucleotide oxidase (methane hydroxylase)
Authors: Ribbons D W
Published: January 2020

Cell-free particulate fractions of extracts from the obligate methylotroph Methylococcus capsulatus catalyze the reduced nicotinamide adenine dinucleotide (NADH) and O2-dependent oxidation of methane (methane hydroxylase). The only oxidation product detected was formate. These preparations also catalyze the oxidation of methanol and formaldehyde to formate in the presence or absence of phenazine methosulphate with oxygen as the terminal electron acceptor. Methane hydroxylase activity cannot be reproducibly obtained from disintegrated cell suspensions even though the whole cells actively respired when methane was presented as a substrate. Varying the disintegration method or extraction medium had no significant effect on the activities obtained. When active particles were obtained, hydroxylase activity was stable at 0 C for days. Methane hydroxylase assays were made by measuring the methane-dependent oxidation of NADH by O2. In separate experiments, methane consumption and the accumulation of formate were also demonstrated. Formate is not oxidized by these particulate fractions. The effects of particle concentration, temperature, pH, and phosphate concentration on enzymic activity are described. Ethane is utilized in the presence of NADH and O2. The stoichiometric relationships of the reaction(s) with methane as substrate were not established since (i) the presumed initial product, methanol, is also oxidized to formate, and (ii) the contribution that NADH oxidase activity makes to the observed consumption of reactants could not be assessed in the presence of methane. Studies with known inhibitors of electron transport systems indicate that the path of electron flow from NADH to oxygen is different for the NADH oxidase, methane hydroxylase, and methanol oxidase activities.