bluuebunny/crossref_metadata_2025 huggingface dataset:
Coupling of Energy to Folate Transport in
Lactobacillus casei
Authors: Henderson Gary B. , Zevely Edward M. , Huennekens F. M.
Published: January 2020

Lactobacillus casei
cells can accumulate folate to an intracellular concentration in excess of 500 μM and to concentration gradients (relative to the extracellular compartment) of several thousand-fold. Maximum rates of folate transport are achieved rapidly (
t
1/2
< 1 min) after the addition of glucose to energy-depleted cells and occur at intracellular adenosine 5'-triphosphate concentrations above 625 μM. The rate of folate transport and the adenosine 5'-triphosphate content of cells are both extremely sensitive to arsenate and decrease in parallel with increasing concentrations of the inhibitor, indicating a requirement for phosphate-bond energy in the transport process. The energy source is not a membrane potential or a pH gradient generated via the membrane-bound adenosine triphosphatase, since dicyclohexylcarbodiimide (an adenosine triphosphatase inhibitor) and carbonyl cyanide
m
-chlorophenylhydrazone (a proton conductor) have little effect on the uptake process. The K
+
-ionophore, valinomycin, is an inhibitor of folate transport, but does not act via a mechanism involving dissipation of the membrane potential. This can be deduced from the facts that the inhibition by valinomycin is relatively insensitive to pH, is considerably greater in Na
+
- than in K
+
-containing buffers, and is not enhanced by the addition of proton conductors. Folate efflux is not affected by valinomycin, glucose, or various metabolic inhibitors, although a rapid release of the accumulated vitamin can be achieved by the addition of unlabeled folate together with an energy source (glucose). These results suggest that the active transport of folate into
L. casei
is energized by adenosine 5'-triphosphate or an equivalent energy-rich compound, and that coupling occurs not via the membrane-bound adenosine triphosphatase but by direct interaction of the energy source with a component of the transport system.
Very slow growth of Escherichia coli
Authors: Chesbro W , Evans T , Eifert R
Published: January 2020

A recycling fermentor (a chemostat with 100% biomass feedback) was used to study glucose-limited behavior of Escherichia coli B. The expectation from mass transfer analysis that growth would asymptotically approach a limit mass determined by the glucose provision rate (GPR) and the culture's maintenance requirement was not met. Instead, growth proceeded at progressively lower rates through three distinct phases. After the fermentor was seeded, but before glucose became limiting, growth followed the usual, exponential path (phase 1). About 12 h postseeding, residual glucose in the fermentor fell below 1 microgram . ml-1 and the growth rate (dx/dt) became constant and a linear function of GPR (phase 2). The specific growth rate, mu, therefore fell continuously throughout the phase. Biomass yield and glucose assimilation (13%) were near the level for exponential growth, however, and independent of GPR over a broad range. At a critical specific growth rate (0.04 h-1 for this strain), phase 2 ended abruptly and phase 3 commenced. In phase 3, the growth rate was again constant, although lower than in phase 2, so that mu continued to fall, but growth rates and yields were praboloid functions of GPR. They were never zero, however, at any positive value of GPR. By inference, the fraction of metabolic energy used for maintenance functions is constant for a given GPR, although different for phases 2 and 3, and independent of biomass. In both phases 2 and 3, orcinol, diphenylamine, and Lowry reactive materials were secreted at near-constant rates such that over 50% as much biosynthetic mass was secreted as was retained by the cells.
Antipain lethality to Escherichia coli: dependence upon cyclic adenosine 3',5'-monophosphate and its receptor protein
Authors: Swenson P A
Published: January 2020

Antipain kills Escherichia coli K-12 cells in an exponential manner beginning 1 h after its addition. Mutant strains, delta cya and crp, which are unable to synthesize cyclic adenosine 3',5'-monophosphate (cAMP) and the cAMP receptor protein, respectively, are not affected. Addition of cAMP (5 mM) to antipain-treated mutant strains causes killing of delta cya cells, but not crp cells. Thus the lethal effect of antipain is dependent upon cAMP and its receptor protein.
Effect of transcription on RecBC- and RecF-mediated recombination within the tryptophan operon of Escherichia coli K-12
Authors: Cosloy S D
Published: January 2020

Recombination between tryptophan gene mutations within the trp operon was determined among transductants for an outside linked cysB marker under conditions of repression and derepression. These studies, carried out with recipient strains utilizing the RecBC or RecF pathway, or a combination of these pathways of recombination, demonstrate that transcription of trp genes as regulated by the trp repressor has no significant effect on RecBC- or RecF-mediated recombination within the trp operon.
Plasmenylethanolamine: growth factor for cholesterol-reducing Eubacterium
Authors: Mott G E , Brinkley A W
Published: January 2020

A plasmalogen, plasmenylethanolamine, is required for in vitro growth of strains of Eubacterium which convert cholesterol to coprostanol. Plasmenylethanolamine was isolated from calf brain by selective saponification of lipid fractions separated by thin-layer or column chromatography. Cholesterol-containing thioglycolate broth plus purified plasmenylethanolamine or its 2-lyso derivative supported growth of Eubacterium ATCC 21408 and a cholesterol-reducing Eubacterium isolated from baboon feces. Plasmenylethanolamine obtained from commercial sources also supported growth of these organisms, but none of a number of other pure lipids would support growth. Metabolism of the alkenyl ether group of plasmenylethanolamine occurred during growth.
Escherichia coli murein-DD-endopeptidase insensitive to beta-lactam antibiotics
Authors: Keck W , Schwarz U
Published: January 2020

A novel endopeptidase degrading the peptide cross-links in sacculi has been isolated from Escherichia coli and purified to homogeneity. The enzyme has a molecular weight of 30,000 and, in contrast to already known enzymes of similar specificity, remains fully active in the presence of beta-lactam antibiotics. In addition, it is exceptional in being inhibited by single-stranded deoxyribonucleic acid and by some polynucleotides. The possible role of the enzyme in cell division is discussed.
Metabolic suppressors of trimethoprim and ultraviolet light sensitivities of Saccharomyces cerevisiae rad6 mutants
Authors: Lawrence C W , Christensen R B
Published: January 2020

Dominant mutations at two newly identified loci, designated SRS1 and SRS2, that metabolically suppress the trimethoprim sensitivity of rad6 and rad18 strains, have been isolated from trimethoprim-resistant mutants arising spontaneously in rad6-1 rad18-2 strains of the yeast Saccharomyces cerevisiae. The SRS2 mutations also efficiently suppress the ultraviolet light sensitivity of the parent strains. They do not, however, suppress their sensitivity to ionizing radiation or their deficiency with respect to induced mutagenesis and sporulation. Such observations support the hypothesis that RAD6-dependent activities can be separated into two functionally distinct groups: a group of error-free repair activities that are responsible for a large amount of the radiation resistance of wild-type strains and also for their resistance to trimethoprim, and a group of error-prone activities that are responsible for induced mutagenesis and are also important in sporulation, but which account at best for only a very small amount of wild-type recovery.
Characterization of molybdenum cofactor from Escherichia coli
Authors: Amy N K , Rajagopalan K V
Published: January 2020

Molybdenum cofactor activity was found in the soluble fraction of cell-free extracts of Escherichia coli grown aerobically in media supplemented with molybdate. Cofactor was detected by its ability to complement the nitrate reductase-deficient mutant of Neurospora crossa, nit-1, resulting in the vitro formation of nitrate reductase activity. Acid treatment of E. coli extracts was not required for release of cofactor activity. Cofactor was able to diffuse through a membrane of nominal 2,000-molecular-weight cutoff and was insensitive to trypsin. The cofactor was associated with a carrier molecule (approximately 40,000 daltons) during gel filtration and sucrose gradient centrifugation, but was easily removed from the carrier by dialysis. The carrier molecule protected the cofactor from inactivation by heat or oxygen. E. coli grown in molybdenum-free media, without and with tungsten, synthesized a metal-free "empty" cofactor and its tungsten analog, respectively, both of which were subsequently activated by the addition of molybdate. Empty and tungsten-containing cofactor complemented the nitrate reductase subunits in the nit-1 extract, forming inactive, but intact, 7.9S nitrate reductase. Addition of molybdate to the enzyme complemented in this manner restored nitrate reductase activity.
Purification and properties of a nicotinamide adenine dinucleotide-linked dehydrogenase that serves an Escherichia coli mutant for glycerol catabolism
Authors: Tang C T , Ruch F E , Lin C C
Published: January 2020

Glycerol:NAD+2-OXIDOREDUCTASE (EC 1.1.1.6) was purified to homogeneity from a mutant of Escherichia coli K12 that uses this enzyme, instead of ATP:glycerol 3-phosphotransferase (EC 2.7.1.30), as the first enzyme for the dissimilation of glycerol. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate shows a subunit of 39,000 daltons. During electrophoresis under nondenaturing conditions, the protein migrates as two bands. These two forms, both of which are enzymatically active, appear to be dimers and octomers of the same subunit. The optimal pH for the oxidation of glycerol is about 10, and that for the reduction of dihydroxyacetone is about 6. Glycerol dehydrogenation is highly activated by NH4+, K+, or Rb+, but strongly inhibited by N-ethylmalemide, 8-hydroxyquinoline, 1,10-phenanthroline, Cu2+, and Ca2+. The enzyme exhibits a broad substrate specificity. In addition to glycerol, it act on 1,2-propanediol and several of its analogs.
Protein-Associated Lipid of
Bacillus stearothermophilus
Authors: Card George L. , Szuba Joan C. , Shimizu Marilyn
Published: January 2020

The composition and patterns of metabolism of phospholipids isolated as part of a lipid-depleted membrane fragment (LDM fragment) and associated with the membrane adenosine triphosphatase complex have been compared with those of the bulk membrane phospholipid. The bulk lipid was extracted from washed membranes with sodium cholate. The LDM fragments, which contained a portion of the electron transport system and the membrane adenosine triphosphatase complex, were purified by chromatography with Sepharose 6B. The LDM fragment preparations contained 0.10 ± 0.02 μmol of lipid phosphorus per mg of protein, compared with 0.54 ± 0.05 μmol of lipid phosphorus per mg of protein for washed membranes. The phospholipid associated with the LDM fragments consisted of 78 ± 4% cardiolipin, 7 ± 1% phosphatidylglycerol, and 15 ± 3% phosphatidylethanolamine. Changes in the total membrane lipid composition (produced by culture conditions) did not alter the phospholipid composition of the LDM fragments. The adenosine triphosphate complex was separated from the other components of the LDM fragments by suspension of the fragments in 1% Triton X-100 and precipitation with antibody specific for the F
1
component of the adenosine triphosphatase complex. The phospholipid isolated with the adenosine triphosphatase complex consisted of 86% cardiolipin, 8% phosphatidylglycerol, and 6% phosphatidylethanolamine. In pulse-chase experiments with
32
P and [2-
3
H]glycerol, the labeling patterns of the phosphatididylglycerol and phosphatidylethanolamine associated with the LDM fragments were different from those of the bulk membrane phosphatidylglycerol and phosphatidylethanolamine. It was concluded that at least a portion of the phospholipid isolated with the LDM fragments was part of a native lipid-protein complex.
Immuno-electron microscopic localization of lipopolysaccharide antigens on ultrathin sections of Salmonella typhimurium
Authors: Takamiya H , Batsford S , Gelderblom H , Vogt A
Published: January 2020

Lipopolysaccharide antigens were demonstrated on ultrathin sections of styrene-embedded Salmonella typhimurium by direct postembedding staining with ferritin-labeled antibodies. The antigenicity, partially masked in the embedding process, could be satisfactorily recovered by treatment of ultrathin sections with nonspecific protease. As judged from the reaction site of the ferritin-labeled antibodies, the lipopolysaccharides were localized in two zones. The broader zone of densely distributed ferritin molecules was superimposed over the whole outer cellwall, and a smaller zone revealing antigenicity was found over the cell membrane, which strongly supports the concept that the latter is the site of synthesis of lipopolysaccharides. The well-defined labeled areas between these two antigenic zones may be the routes whereby the synthesized polysaccharide molecules reach the cell wall.
Role of the ftsA gene product in control of Escherichia coli cell division
Authors: Donachie W D , Begg K J , Lutkenhaus J F , Salmond G P , Martinez-Salas E , Vincente M
Published: January 2020

The kinetics of cell division have been studied in a strain of Escherichia coli which has an amber mutation in the ftsA gene and which also carries a temperature sensitive amber suppressor. This strain is therefore temperature sensitive for the synthesis of the ftsA protein. Cells of this strain were able to divide only if the synthesis of this protein took place during a specific part of the cell cycle. This was a short period (roughly 10 min in duration) immediately before the normal time of cell division.
Function of Modified Nucleosides 7-Methylguanosine, Ribothymidine, and 2-Thiomethyl-
N
6
-(Isopentenyl)adenosine in Procaryotic Transfer Ribonucleic Acid
Authors: Hoburg A. , Aschhoff H. J. , Kersten H. , Manderschied U. , Gassen H. G.
Published: January 2020

To elucidate subtle functions of transfer ribonucleic acid (tRNA) modifications in protein synthesis, pairs of tRNA's that differ in modifications at specific positions were prepared from
Bacillus subtilis
. The tRNA's differ in modifications in the anticodon loop, the extra arm, and the T C loop. The functional properties of these species were compared in aminoacylation, as well as in initiation and peptide bond formation, at programmed ribosomes. These experiments demonstrated the following. (i) In tRNA
f
Met
the methylation of guanosine 46 in the extra arm to 7-methylguanosine by the 7-methylguanosine–forming enzyme from
Escherichia coli
changes the aminoacylation kinetics for the
B. subtilis
methionyl-tRNA synthetase. In repeated experiments the
V
max
value is decreased by one-half. (ii) tRNA
f
Met
species with ribothymidine at position 54 (rT54) or uridine at position 54 (U54) were obtained from untreated or trimethoprim-treated
B. subtilis
. The formylated fMet-tRNA
f
Met
species with U54 and rT54, respectively, function equally well in an in vitro initiation system containing AUG, initiation factors, and 70s ribosomes. The unformylated Met-tRNA
t
Met
species, however, differ from each other: “Met-tRNA
f
Met
rT” is inactive, whereas the U54 counter-upart effectively forms the initiation complex. (iii) Two isoacceptors, tRNA
1
Phe
and tRNA
2
Phe
, were obtained from
B. subtilis
. tRNA
1
Phe
accumulates only under special growth conditions and is an incompletely modified precursor oftRNA
2
Phe
: in the first position of the anticodon, guanosine replaces Gm, and next to the 3′ end of the anticodon (isopentenyl)adenosine replaces 2-thiomethyl-
N
6
-(isopentenyl)adenosine. Both tRNA's behave identically in aminoacylation kinetics. In the factor-dependent AUGU
3
-directed formation of fMet-Phe, the undermodified tRNA
1
Phe
is always less efficient at Mg
2+
concentrations between 5 and 15 mM than its mature counterpart.
Carbon dioxide assimilation in cyanobacteria: regulation of ribulose, 1,5-bisphosphate carboxylase
Authors: Tabita F R , Colletti C
Published: January 2020

Cyanobacteria assimilate carbon dioxide through the Calvin cycle and therefore must regulate the activity of ribulose 1,5-bisophosphate carboxylase. Using an in situ assay, as well as measuring the activity in crude, partially purified, and homogeneous preparations, we can show that a number of phosphorylated intermediates exert a regulatory role. Three diverse organisms, Agmenellum quadruplicatum, Aphanocapsa 6714, and Anabaena sp. CA, were studied, and it was found that the in situ and cell-free carboxylase activities were particularly affected by low levels of phosphogluconate and reduced nicotinamide adenine dinucleotide phosphate. There was a marked activation by these ligands when the inactive enzyme was assayed in the presence of low levels of bicarbonate, a result significantly different from a previous report. Moreover, the fully activated enzyme was inhibited by phosphogluconate. In situ Anabaena CA carboxylase activity exhibited a particular capacity for activation by phosphogluconate and reduced nicotinamide adenine dinucleotide phosphate. However, activation of the crude, partially purified, or homogeneous Anabaena CA carboxylase by phosphogluconate and reduced nicotinamide adenine dinucleotide phosphate was significantly decreased when compared with enzyme activity in permeabilized cells. It appears that the microenvironment or the conformation of the enzyme within the cell may be significantly different from that of the isolated enzyme.
Defective F pili and other characteristics of Flac and Hfr Escherichia coli mutants resistant to bacteriophage R17
Authors: Burke J M , Novotny C P , Fives-Taylor P
Published: January 2020

Mutants resistant to the donor-specific bacteriophage R17 were isolated from Hfr and Flac-containing strains of Escherichia coli K-12. Thirty-five mutants were examined for the presence of F pili by electron microscopy. The pilus morphology was studied, as were the abilities of the cells to retract their pili and to synthesize new pili. Measurements were made of the efficiency of the conjugal deoxyribonucleic acid transfer and of M13 and R17 phage infection. All mutants had noticeable defects in pilus production, structure, or function. Mutants were found which produced unusually long pili, displayed wide variations in the number of pili per cell, and were deficient in pilus retraction and synthesis. Evidence is presented that there may be two pathways of pilus retraction.
Protein turnover in the extreme thermophile Thermus aquaticus
Authors: Kenkel T , Trela J M
Published: January 2020

Protein turnover in the extreme bacterial thermophile Thermus aquaticus was examined in exponential cultures at 75 degrees C. The relative amount of [3H]leucine incorporated into trichloroacetic acid-insoluble material was stable in pulse-chase experiments assayed over 2.5 h. The trichloroacetic acid-insoluble radioactive leucine was stable upon the addition of chloramphenicol, which blocks protein synthesis in T. aquaticus. The specific activity of a phosphate-repressible alkaline phosphatase, investigated in the presence of chloramphenicol, did not decrease. The addition of excess orthophosphate to cultures derepressed for the alkaline phosphatase did not show a marked effect on the specific activity over a 2-h period. On the basis of these four experiments, it does not appear that a high protein turnover rate is essential for the thermophily of T. aquaticus at 75 degrees C.
Deoxyribonucleic Acid Sequence Homologies Among Bacterial Insertion Sequence Elements and Genomes of Various Organisms
Authors: Nisen Perry , Purucker Mary , Shapiro Lucille
Published: January 2020

Plasmid and phage deoxyribonucleic acid (DNA) harboring bacterial insertion sequence (IS) elements IS
1
, IS
2
, and IS
5
were characterized and used as probes to detect homologous sequences in various procaryotic and eucaryotic genomes. The hybridization method used permits the detection of sequences partially homologous to the elements. Hybridization of the IS-containing probes to each other revealed a region of limited homology between IS
1
and IS
2
. Homologous sequences were then detected by computer analysis of the published IS
1
and IS
2
nucleotide sequences. The homologous sequence contains a tandemly repeated tetranucleotide sequence which resembles the repeated sequence at the hot spot for spontaneous mutations in the
lacI
gene (P. J. Farabaugh, U. Schmeissner, M. Hofer, and J. Miller, J. Mol. Biol.
126:
847–863, 1978). Homology between the IS elements and various genomes was determined by hybridizing labeled DNA containing IS
1
, IS
2
, and IS
5
sequences to Southern blots of chromosomal DNA cleaved with restriction endonucleases. IS
1
and IS
5
appear limited to the enteric bacteria, whereas IS
2
sequences can also be detected in
Pseudomonas putida, Pseudomonas aeruginosa
, and
Serratia marcescens
. Bacteria which appear not to possess extrachromosomal elements, e.g.,
Caulobacter crescentus
, did not show homology with any insertion sequences tested. In addition, sequences homologous to IS
1
, IS
2
, or IS
5
were not detected in
Saccharomyces cerevisiae, Dictyostelium discoideum
, or calf thymus DNA.
Completed Bacillus subtilis nucleoid as a doublet structure
Authors: McGinness T , Wake R G
Published: January 2020

When outgrowing spores of the temperature-sensitive dna initiation mutants of Bacillus subtilis, TsB134 and dna-1, were allowed to undergo a single round of replication by shifting to the restrictive temperature soon after its initiation, both segregating daughter nucleoids appeared as clearly defined doublet structures. The components of each doublet remained together as a discrete pair, even under conditions which resulted in the formation of deoxyribonucleic acid (DNA)-less cells. A doublet nucleoid was also observed at a high frequency when TsB134 spores were allowed to germinate and grow out in the complete absence of DNA synthesis at the permissive temperature. TsB134 spores were foud to contain the usual "haploid" amount of DNA. It is suggested that the doublet nucleoid reflects a folding of a single chromosome into two large domains which resolve from one another under conditions of cell extension in the absence of DNA synthesis.
Amphilphilic nature of spiralin, the major protein of the Spiroplasma citri cell membrane
Authors: Wróblewski H
Published: January 2020

Spiralin could not be solubilized in the absence of detergents, and it was shown by charge-shift crossed immunoelectrophoresis that this protein was capable of binding detergents under nondenaturing conditions. These properties indicate the amphiphilic nature of spiralin, which therefore should be regarded as an intrinsic membrane protein. The efficiency of mild (ionic and neutral) detergents to solubilize spiralin was as follows: deoxycholate greater than lauroyl sarcosinate, cholate, taurocholate, taurodeoxycholate greater than Triton X-100 greater than Brij 58 greater than Tween 20, indicating that mild ionic detergents were more effective than neutral ones. Solubilization of spiralin was quantitative with sodium deoxycholate. It was also shown that although a membrane protein is not extractable by a given detergent from the membrane, this does not necessarily mean that the protein is not soluble in this detergent.
Cobalamins and cobalamin-dependent enzymes in Candida utilis
Authors: Poston J M , Hemmings B A
Published: January 2020

Candida utilis has been shown to contain 4.7 pmol of cobalamin per g of wet cell paste. Purification of the cobalamin showed it to be a mixture of methylcobalamin and adenosylcobalamin. Two cobalamin-dependent enzyme systems have been found in the yeast: methylcobalamin-dependent methionine biosynthesis and leucine 2,3-aminomutase. The cobalamin extracted from the yeast is as effective as authentic adenosylcobalamin in stimulating leucine 2,3-aminomutase.
Regulation of membrane peptides by the Pseudomonas plasmid alk regulon
Authors: Benson S , Oppici M , Shapiro J , Fennewald M
Published: January 2020

Pseudomonas putida strains carrying the plasmid alk genes will grow on n-alkanes. Induced alk+ strains contain membrane activities for alkane hydroxylation and dehydrogenation of aliphatic primary alcohols. P. putida cytoplasmic and outer membranes can be separated by sucrose gradient centrifugation after disruption of cells by either mild detergent lysis or passage through a French press. Both the membrane component of alkane hydroxylase and membrane alcohol dehydrogenase fractionated with the cytoplasmic membrane. Induction of the alk regulon resulted in the appearance of at least three new plasmid-determined cytoplasmic membrane peptides of about 59,000 (59K), 47,000 (47K), and 40,000 (40K) daltons as well as the disappearance of a pair of chromosomally encoded outer membrane peptides of about 43,000 daltons. The 40K peptide is the membrane component of alkane hydroxylase and the product of the plasmid alkB gene because the alkB1029 mutation altered the properties of alkane hydroxylase in whole cells, reduced its thermal stability in cell extracts, and led to increased electrophoretic mobility of the inducible 40K peptide. These results are consistent with a model for vectorial oxidation of n-alkanes in the cytoplasmic membrane of P. putida.
Identification of cistrons involved in conjugal transfer of narrow-host-range R plasmid R91-5 of Pseudomonas aeruginosa
Authors: Carrigan J M , Krishnapillai V
Published: January 2020

The development of a transductional method for complementation tests between transfer-deficient mutants of the narrow-host-range R plasmic R91-5 of Pseudomonas aeruginosa has allowed the indentification of cistrons involved in the conjugal transfer of this plasmid. Complementation tests performed between transfer-deficient mutants characterized phenotypically with respect to sensitivity to donor-specific phage, ability to inhibit the replication of phage G101, and expression of entry-exclusion has identified a minimum of 10 transfer cistrons. Although most mutagen-induced mutants were relatively heterogeneous and appeared to be affected in a single cistron only, a high proportion of mutants isolated after selection for donor-specific phage resistance had deletions but always included tra Y. Mutants selected directly on the basis of transfer deficiency which also became donor-specific phage resistant fell into all 10 cistrons, suggesting that many R91-5 transfer cistrons are concerned with the synthesis of sex pili and other surface structures necessary for conjugal transfer. Conversely, most retaining donor-specific phage sensitivity belonged to one cistron, whereas transfer-deficient mutants which had also lost the ability to inhibit the replication of phage G101 comprised four cistrons.
Outer membrane of Pseudomonas aeruginosa: heat- 2-mercaptoethanol-modifiable proteins
Authors: Hancock R E , Carey A M
Published: January 2020

A number of polyacrylamide gel systems and solubilization procedures were studied to define the number and nature of "major" polypeptide bands in the outer membrane of Pseudomonas aeruginosa. It was shown that five of the eight major outer membrane proteins were "heat modifiable" in that their mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis was determined by the solubilization temperature. Four of these heat-modifiable proteins had characteristics similar to protein II of the Escherichia coli outer membrane. Addition of lipopolysaccharide subsequent to solubilization caused reversal of the heat modification. The other heat-modifiable protein, the porin protein F, was unusually stable to sodium dodecyl sulfate. Long periods of boiling in sodium dodecyl sulfate were required to cause conversion to the heat-modified form. This was demonstrated both with outer membrane-associated and purified lipopolysaccharide-depleted protein F. Furthermore, lipopolysaccharide treatment had no effect on the mobility of heat-modified protein F. Thus it is concluded that protein F represents a new class of heat-modifiable protein. It was further demonstrated that the electrophoretic mobility of protein F was modified by 2-mercaptoethanol and that the 2-mercaptoethanol and heat modification of mobility were independent of one another. The optimal conditions for the examination of the outer membrane proteins of P. aeruginosa by one-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis are discussed.
Membrane-bound respiratory chain of Pseudomonas aeruginosa grown aerobically
Authors: Matsushita K , Yamada M , Shinagawa E , Adachi O , Ameyama M
Published: January 2020

The electron transport chain of the gram-negative bacterium Pseudomonas aeruginosa, grown aerobically, contained a number of primary dehydrogenases and respiratory components (soluble flavin, bound flavin, coenzyme Q9, heme b, heme c, and cytochrome o) in membrane particles of the organism. Cytochrome o, about 50% of the b-type cytochrome, seemed to function as a terminal oxidase in the respiratory chain. The electron transport chain of P. aeruginosa grown aerobically was suggested to be lined up in order of primary dehydrogenase, b, c1, c, o, and oxygen.
Cyanophycin granule polypeptide formation and degradation in the cyanobacterium Aphanocapsa 6308
Authors: Allen M M , Hutchison F , Weathers P J
Published: January 2020

The effect of a number of conditions on the amount of cyanophycin granule polypeptide [multi-L-arginyl poly(L-aspartic acid)] formed in the unicellular cyanobacterium Aphanocapsa 6308 was determined. Light, CO2, sulfur, and phosphorus starvation as well as the addition of arginine to culture media increased the amount of cyanophycin granule polypeptide in cells when compared with that in cells grown under conditions optimal for growth. Nitrogen limitation and reduction of growth temperature to 30 degrees C decreased the amount of cyanophycin granule polypeptide on a dry-weight basis. Shift-up and shift-down experiments suggest cyanophycin granule polypeptide may be a reserve nitrogen polymer in Aphanocapsa 6308.
Origin of hydrogen in methane produced by Methanobacterium thermoautotrophicum
Authors: Daniels L , Fulton G , Spencer R W , Orme-Johnson W H
Published: January 2020

The production of deuterated methane by Methanobacterium thermoautotrophicum in H2O-D2O mixtures was examined by high-resolution mass spectrometry. The hydrogen in the methane arose solely from water and not from hydrogen gas. Hydrogen gas served only as an electron source in methanogenesis. A whole-cell product isotope discrimination of 1.5 favoring hydrogen over deuterium was observed in methane production in 81 atom% deuterated water. The distribution of deuterated methane species is described by a simple model of the overall reaction.
Regulation of Klebsiella pneumoniae hut operons by oxygen
Authors: Goldberg R B , Hanau R
Published: January 2020

We investigated the regulation of genes concerned with nitrogen metabolism by oxygen in the facultative anaerobe Klebsiella pneumoniae. We found oxygen to be required for the expression of the hut operons; the effect of O2 on the glutamine synthetase and urease was less pronounced than on the hut operons. Glutamine synthetase was transiently repressed during the transition from an aerobic to an anaerobic environment. Regulation of hut by O2 suppressed the effect of nitrogen limitation on the expression of these genes.
Mutation in the crp gene of Salmonella typhimurium which interferes with inducer exclusion
Authors: Scholte B J , Postma P W
Published: January 2020

A mutation in the crp gene of Salmonella typhimurium is described which overcame the defects of ptsHI deletion mutants for growth on a number of nonphosphotransferase system compounds. This mutation abolished inducer exclusion in a leaky ptsI mutant. The possible implications for the mechanism of inducer exclusion are discussed.
Localization of enterobacterial common antigen: Proteus mirabilis and its various L-forms
Authors: Rinno J , Gmeiner J , Golecki J R , Mayer H
Published: January 2020

An investigation of Proteus mirabilis wild-type strains and their various derived L-forms shows that the enterobacterial common antigen (ECA) is localized in the outer membrane of the cell envelope of these strains. In strains where the outer membrane is lacking (stable protoplast L-forms) or where its amount is reduced (spheroplast UL19) no ECA or only reduced amounts of it are detected by serological tests or by ferritin-labeling techniques.
Dictyostelium discoideum fruiting bodies observed by scanning electron microscopy
Authors: Murata Y , Ohnishi T
Published: January 2020

The fruiting bodies of Dictyostelium discoideum, fixed in a vapor of 25% glutaraldehyde, were observed by scanning electron microscopy. The surface of the basal disk had rootlike ripples, sometimes with concentric rings of riblike striations at its periphery. The surface of the stalk had pleated and twisted ridges along its entire length, but no lateral projections. The spores were randomly orientated and had a crinkled surface. The surface of the lower part of the papilla had flattened ridges running horizontally. A slime coat remained in the region of transition between the stalk and sporangium, but not on the spores.
Structure and composition of cyanophycin granules in the cyanobacterium Aphanocapsa 6308
Authors: Allen M M , Weathers P J
Published: January 2020

Cyanophycin granules in the unicellular cyanobacterium Aphanocapsa 6308 were examined with the electron microscope in both thin section and by freeze-fracture techniques. Purified granules were examined with the electron microscope, by arginine determinations, by chromatography, and by elemental analysis. They are similar in ultrastructure and composition to those isolated from the nitrogen-fixing cyanobacterium Anabaena cylindrica, consisting of equal molar quantities of L-arginine and L-aspartic acid.
Visualization of cell-cell contact during conjugation in Streptococcus faecalis
Authors: Krogstad D J , Smith R M , Moellering R C , Parquette A R
Published: January 2020

Conjugal transfer of an antibiotic resistance plasmid in Streptococcus faecalis was associated with direct intercellular connections between chains of streptococci in the absence of either pili or fimbriae.
Induction of synchronous growth in the yeast phase of Wangiella dermatitidis
Authors: Roberts R L , Lo R J , Szaniszlo P J
Published: January 2020

Synchronous yeast-phase cultures of Wangiella dermatitidis were induced by starvation, heat shock, and inhibition of deoxyribonucleic acid synthesis by hydroxyurea. Hydroxyurea-induced synchrony resulted in some distortion of the yeast-phase cell cycle. However, induction of synchrony by hydroxyurea is a rapid and simple technique which generates a marked degree of synchronous growth.
Selection of Bacillus subtilis mutants impaired in ammonia assimilation
Authors: Dean D R , Aronson A I
Published: January 2020

The selection of Bacillus subtilis mutants capable of using D-histidine to fulfill a requirement for L-histidine resulted in mutants with either no glutamate synthase activity or increased amounts of an altered glutamine synthetase.
Composition of cardiolipin molecular species in Escherichia coli
Authors: Yokota K , Kanamoto R , Kito M
Published: January 2020

The composition of the molecular species of acidic phospholipids in Escherichia coli B during the late exponential growth phase at 37 degrees C was determined. Two phosphatidyl groups of cardiolipin, the 3-(3-sn-phosphatidyl) and 1-(3-sn-phosphatidyl) moieties of cardiolipin, were isolated by limited hydrolysis with phospholipase C. No significant difference in the composition of the molecular species was found between the 3-(3-sn-phosphatidyl) and 1-(3-sn-phosphatidyl) moieties. On the other hand, the composition of the molecular species of phosphatidylglycerol was different from that of cardiolipin. Phosphatidylglycerol contained more of the 1-palmitoyl 2-cis-9,10-methylenehexadecanoyl and 1-palmitoyl 2-cis-11,12-methyleneoctadecanoyl species than did cardiolipin. The difference in the composition of the molecular species between cardiolipin and phosphatidylglycerol may depend on the difference in the turnover rates of both phospholipids.
Hairy root: plasmid encodes virulence traits in Agrobacterium rhizogenes
Authors: White F F , Nester E W
Published: January 2020

Agrobacterium rhizogenes strain 15834, which incites hairy root disease in plants, harbors three large plasmids: pAr15834a (107 x 10(6) daltons), pAr15834b (154 x 10(6) daltons), and pAr15834c (258 x 10(6) daltons). Kanamycin-resistant transconjugants were selected in a cross of kanamycin-resistant derivate of strain 15834 and an avirulent recipient. The transconjugants belonging to one class were virulent and contained all three donor plasmids. These transconjugants also acquired sensitivity to the bacteriocin agrocin 84. The loss of plasmids from virulent transconjugants during growth at 37 degrees C indicated that virulence genes reside on pAr15834b, whereas agrocin 84 sensitivity genes reside on pAr15834a. The pathology induced by the virulent transconjugants containing only pAr15834b was identical to that produced by the wild-type strain of A. rhizogenes. Restriction endonuclease fragment analysis of plasmids from the transconjugants and the donor revealed that pAr15834c is a cointegrate of pAr15834a and pAr15834b. Kanamycin-resistant transconjugants belonging to a second class were avirulent and contained an altered form of pAr15834b. Strain 15834 can utilize octopine. However, this trait was not detected in any of the transconjugants. Octopine is not synthesized by infected plant tissue.
Mapping of insertion mutations in gnd of Escherichia coli with deletions defining the ends of the gene
Authors: Wolf R E , Cool J A
Published: January 2020

A genetic map was prepared for gnd, the gene of Escherichia coli which encodes the metabolically regulated 6-phosphogluconate dehydrogenase. Direct selection methods were used for the isolation of mutants with deletions that define the respective ends of gnd. These selections depended on the availability of a defective lysogen in which gnd was present on a lambda h80 dgnd his prophage located at the att phi 80 region of the chromosome. Mutants with deletions entering gnd from the his-distal end were selected as Gnd- TonB- mutants. Mutants with his-proximal gnd deletions were selected as Gnd-, temperature-resistant mutants of a specially prepared stable lysogen. Gnd- mutants were also isolated after mutagenesis with bacteriophage Mu cts61, and genetic tests were used to determine which mutants carry a Mu cts61 prophage in gnd. The deletion mutations were mapped against each other and against the insertion mutations through the use of F' merodiploid strains. The insertion mutations mapped at seven distinct sites in gnd; three mapped under the deletions defining the his-proximal portion of the gene and three mapped with the his-distal deletions.
Characterization of the cell wall and cell wall proteins of Chromatium vinosum
Authors: Lane B C , Hurlbert R E
Published: January 2020

Highly purified cell walls of Chromatium vinosum were isolated by differential centrifugation, with or without Triton X-100 extraction. The isolated material had a protein composition similar to that of cell walls obtained by sucrose density gradient centrifugation. Twenty-two proteins were reproducibly detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A 42-kilodalton protein was shown to account for 65% of the total cell wall protein. The majority of cell wall proteins were solubilized in sodium dodecyl sulfate at room temperature; however, they existed as high-molecular-weight complexes unless heated to 45 degrees C or above. The cell wall contained one heat-modifiable protein which migrated with an apparent molecular weight of 37,400 when solubilized at 70 degrees C or below, but which migrated with an apparent molecular weight of 52,500 if solubilized at 100 degrees C. The electrophoretic mobility of three proteins was modified by 2-mercaptoethanol. The majority of C. vinosum cell wall proteins had isoelectric points between pH 4.5 and 5.5, and the 42-kilodalton protein focused at pH 4.9. No proteins were detected which were analogous to the lipoprotein or peptidoglycan-associated proteins of the Enterobacteriaceae. Nearest-neighbor analysis with a reducible, cross-linking reagent indicated that three proteins, including the 42-kilodalton protein, associated with themselves. Most of the cell wall proteins were partially accessible to proteases in both intact cells and isolated cell walls. Protease treatment of the whole cell or isolated cell wall digested approximately an 11,000-molecular-weight portion of the 42-kilodalton protein.
Acceptance by
Erwinia
spp. of R Plasmid R68.45 and Its Ability to Mobilize the Chromosome of
Erwinia chrysanthemi
Authors: Chatterjee Arun K.
Published: January 2020

R plasmid R68.45 was transferred in broth matings from
Escherichia coli
to strains of
Erwinia amylovora, E. carotovora
subsp.
atroseptica, E. chrysanthemi
, and
E. herbicola
(
Enterobacter agglomerans
); the frequency of transfer ranged from 2 × 10
−8
to 5 × 10
−4
per input donor cell depending on the bacterial species. The drug resistance markers
tet
+
,
amp
+
, and
kan
+
were stable in these
Erwinia
species. Transconjugants of
Erwinia
spp., but not of the wild-type parent
Erwinia
strains, acquired levels of antibiotic resistance (tetracycline, 50 μg/ml; ampicillin, 200 μg/ml; kanamycin 200 μg/ml) similar to those of the donor R68.45-bearing strain of
Escherichia coli. Erwinia
transconjugants (with one exception of
E. carotovora
subsp.
atroseptica
) were donors of the antibiotic resistance markers; the frequency of transfer was consistently higher with an
E. coli
strain than with
Erwinia
spp. as recipients, and when matings were done on a solid surface (membranes) rather than in liquid. Transfer of chromosomal markers
ade
+
,
gal
+
,
gtu
+
(utilization of galacturonate),
his
+
,
leu
+
,
lys
+
,
thr
+
, and
trp
+
occurred in crosses between
E. chrysanthemi
strains harboring R68.45 and appropriate recipient strains; the frequency of transfer ranged from 9.0 × 10
−8
to 2.0 × 10
−6
depending on the selective marker. Analysis of the coinheritance of unselected markers among various classes of recombinants revealed linkage between
thr-leu-lys-ade
and between
trp
and
his
, thus confirming earlier findings with the Hfr-type donor cells. Since R68.45 mobilized an array of chromosomal markers in the wild-type as well as genetically marked strains of
E. chrysanthemi
, the system, used in conjunction with the existing Hfr strains, should provide a useful tool to study the genetics of plant pathogenicity of this bacterial species. In contrast to
E. chrysanthemi
, R68.45 did not mobilize chromosomal markers
ilv
+
,
his
+
,
rbs
+
,
ser
+
, and
thr
+
in
E. amylovora
EA178.
Hydrogenase in actinorhizal root nodules and root nodule homogenates
Authors: Benson D R , Arp D J , Burris R H
Published: January 2020

Hydrogenases were measured in intact actinorhizal root nodules and from disrupted nodules of Alnus glutinosa, Alnus rhombifolia, Alnus rubra, and Myrica pensylvanica. Whole nodules took up H2 in an O2-dependent reaction. Endophyte preparations oxidized H2 through the oxyhydrogen reaction, but rates were enhanced when hydrogen uptake was coupled to artificial electron acceptors. Oxygen inhibited artifical acceptor-dependent H2 uptake. The hydrogenase system from M. pensylvanica had a different pattern of coupling to various electron acceptors than the hydrogenase systems from the alders; only the bayberry system evolved H2 from reduced viologen dyes.
Regulation of enzymes of the 3,5-xylenol-degradative pathway in Pseudomonas putida: evidence for a plasmid
Authors: Hopper D J , Kemp P D
Published: January 2020

Constitutive synthesis of enzymes responsible for methyl group oxidation in 3,5-xylenol degradation and an associated p-cresol methylhydroxylase in Pseudomonas putida NCIB 9869 was shown by their retention at high specific activities in cells transferred from 3,5-xylenol medium to glutamate medium. The specific activities of other enzymes of the 3,5-xylenol pathway declined upon removal of aromatic substrate, consistent with their inducible control. Specific activities of the methyl-oxidizing enzymes showed an eventual decline concomitant with a decrease in the fraction of bacteria capable of growth with 3,5-xylenol; a simultaneous loss of the ability to grow with m-hydroxybenzoate was also observed. The property of 3,5-xylenol utilization could be transferred to another strain of P. putida. It is proposed that enzymes of the 3,5-xylenol pathway and those for conversion of p-cresol to p-hydroxybenzoate are plasmid encoded, that the early methyl-oxidizing enzymes are expressed constitutively, and that the later enzymes are inducible.
Peptone Induction and Rifampin-Insensitive Collagenase Production by
Vibrio alginolyticus
Authors: Reid Graham C. , Woods David R. , Robb Frank T.
Published: January 2020

Vibrio alginolyticus
produces an extracellular collagenase which requires specific induction by collagen or its high-molecular-weight fragments. Peptone also induces collagenase during the late exponential and early stationary growth phases. The peptone inducers have been shown to have a broad molecular weight range between 1,000 and 60,000. The peptone inducers supported slow growth of
V. alginolyticus
when supplied as the sole nitrogen source in minimal medium. Digestion of the peptone inducers with purified
V. alginolyticus
collagenase resulted in a decrease in their inducing ability, whereas digestion with trypsin or α-chymotrypsin did not. This indicated that induction by the inducers required the presence of collagenase-sensitive bonds. Prolonged digestion of the inducers with collagenase did not completely eliminate the inducing ability of the inducers. The peptone inducers acted as inhibitors of collagenase. A minimal medium induction system has been developed which involves resuspending cells at high density in a medium containing succinate, (NH
4
)
2
SO
4
, KH
2
PO
4
, and the peptone inducer. Cells grown in minimal medium induce earlier than cells grown on peptone, Casamino Acids, or tryptone. Collagenase production was shown to occur for 30 to 60 min in the presence of rifampin at levels which completely inhibit the incorporation of [
3
H]uracil into trichloroacetic acid-precipitable material. Chloramphenicol completely and immediately abolished collagenase production, which together with labeling studies has confirmed that collagenase production involves de novo synthesis of the enzyme. Both glucose and Casamino Acids repressed collagenase production, although synthesis of the enzyme continued for 30 to 60 min after their addition. The repression of collagenase production by glucose and Casamino Acids was more severe than the inhibition of enzyme formation due to addition of rifampin.