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E W Nester

Publications and source records attributed to E W Nester.

At least 163 records · Page 9Linked to original sources
PubMed1971

Control of tryptophan biosynthesis by the methyltryptophan resistance gene in Bacillus subtilis.

5-Methyltryptophan-resistant mutants derived from Bacillus subtilis strain 168 synthesize all of the tryptophan biosynthetic enzymes constitutively and excrete tryptophan. These mutants can be divided into three classes: class 1, low enzyme level and low rate of tryptophan excretion; class 2, high enzyme level and intermediate rate of tryptophan excretion; and class 3, high enzyme level and high rate of tryptophan excretion. A bradytrophic requirement for phenylalanine is correlated with the rate of tryptophan excretion. The phenylalanine requirement is relieved when the rate of tryptophan excretion is reduced by either (i) lowering the level of the tryptophan enzymes, (ii) reducing the supply of a tryptophan precursor (chorismate), or (iii) stopping tryptophan synthesis by a mutational block in the pathway. All of the mutants map in a region of the chromosome previously reported as the mtr locus. Our data show that synthesis of the tryptophan enzymes is controlled through the mtr locus but not influenced by precursors of tryptophan.

Aldehyde-Lyases↗
PubMed1971

Macromolecular synthesis in Bacillus subtilis during development of the competent state.

Rates of deoxyribonucleic acid, ribonucleic acid, and protein synthesis were examined in purified competent cells of Bacillus subtilis during the development of the transformable state. To become competent, a cell must depart from the normal course of vegetative growth and pass through a precompetent phase beginning as early as 90 to 180 min before the appearance of transformability. While in the precompetent state, the cell decreases its rate of deoxyribonucleic acid synthesis and lowers its ratio of ribonucleic acid synthesis to protein synthesis. This altered pattern of synthesis eventually leads to a decreased buoyant density of precompetent cells. Once a cell has become both precompetent and low in density, it can be converted to a competent (transformable) cell. The early alterations in macromolecular synthesis were found in two competence regimens, one utilizing a nutritional step-down and one free of such a shift. The data imply that the precompetent state is a generalized characteristic of the B. subtilis transformation system and is not specific to the procedure used to allow competence development. Since precompetence-specific events occur very early in a competence regimen, we conclude that the induction of precompetence is unrelated to sporulation or a nutritional shift.

Bacillus subtilis↗
PubMed1970

Possible inhibitory effect of teichoic acid on Bacillus subtilis transfer ribonucleic acid.

1. tRNA of Bacillus subtilis was found to be variably contaminated with membrane teichoic acid. 2. Samples with high contents of teichoic acid showed no accepting activity for tRNA(Phe) and tRNA(Tyr). 3. Removal of teichoic acid restored accepting activity and fractions containing teichoic acid, separated on Sephadex G-150, inhibited the charging of tRNA(Tyr). 4. The presence of teichoic acid did not inhibit the charging of tRNA(His).

Bacillus subtilis↗
PubMed1969

Mutants of Escherichia coli with an altered tyrosyl-transfer ribonucleic acid synthetase.

We have isolated several mutants defective in the gene for tyrosyl-transfer ribonucleic acid (tRNA) synthetase (tyrS). One of these mutants is described in detail. It was isolated as a tyrosine auxotroph with defects both in the tyrosyl-tRNA synthetase and in the tyrosine biosynthetic enzyme, prephenate dehydrogenase. It also had derepressed levels of the tyrosine-specific 3-deoxy-d-arabinoheptulosonic acid-7-phosphate (DAHP) synthetase. The latter finding suggested that a wild-type tyrS gene was required for repression of the tyrosine biosynthetic enzymes. The following results demonstrated that this hypothesis was not correct. (i) When the defective tyrS gene was transferred to another strain, the tyrosine-specific DAHP synthetase in that strain was not derepressed, and (ii) two other mutants with defective tyrosyl-tRNA synthetases had repressed levels of the tyrosine biosynthetic enzymes. The tyrS gene was located near minute 32 on the Escherichia coli chromosome by interrupted mating experiments.

Amino Acids↗
PubMed1969

Heat-sensitive step in deoxyribonucleic acid-mediated transformation of Bacillus subtilis.

Over 90% of the competent cells in a population of Bacillus subtilis lost their competence after being heated to 50 C for 5 min. There was only a slight loss in the number of transformants if the culture was heated for 5 min after the termination of transformation, but 90% of the transformants were lost after 1 hr at 50 C. The population as a whole grew at a slightly faster rate at 50 C than at 32 C. We postulate that a heat-labile factor is required for the uptake or retention (or both) of deoxyribonucleic acid (DNA) in the cell, since uptake of (32)P-DNA into a deoxyribonuclease-resistant form was inversely proportional to the time of exposure to heat. Cells that had lost competence after being heated did not regain their competence for at least several hours, although other cells in the population became competent. These data suggest that the heat-labile factor required for competence is synthesized only once during the period that a cell remains competent.

Bacillus subtilis↗
PubMed1969

Regulation of enzyme synthesis in the aromatic amino acid pathway of Bacillus subtilus.

The control of the synthesis of certain key enzymes of aromatic amino acid biosynthesis was studied. Tyrosine represses the first enzyme of the 3-deoxy-d-arabino heptulosonic acid 7-phosphate pathway, DAHP synthetase, as well as shikimate kinase and chorismate mutase about fivefold in cultures grown under conditions limiting the synthesis of the aromatic amino acids. A mixture of tyrosine and phenylalanine represses twofold further. Tryptophan does not appear to be involved in the control of these enzymes. The specific activity of at least one early enzyme, dehydroquinase, remains essentially constant under a variety of nutritional supplementations. Two enzymes in the terminal branches are repressed by the amino acids they help to synthesize: prephenate dehydrogenase can be repressed fourfold by tyrosine, and anthranilate synthetase can be repressed over 200-fold by tryptophan. There is no evidence that phenylalanine represses prephenate dehydratase. Regulatory mutants have been isolated in which various enzymes of the pathway are no longer repressible. One class is derepressed for several of the prechorismate enzymes, as well as chorismate mutase and prephenate dehydrogenase. In another mutant, several enzymes of tryptophan biosynthesis are no longer repressible. Thus, the rate of synthesis of enzymes at every stage of the pathway is under control of various aromatic amino acids. Tyrosine and phenylalanine control the synthesis of enzymes involved in the synthesis of the three aromatic amino acids. Each terminal branch is under the control of its end product.

Arabinose↗
PubMed1969

Heat-activated endonuclease in Bacillus subtilis.

A heat-stable, heat-activated endonuclease was found in sonic extracts of the transformable strain Bacillus subtilis 168 when the organism was grown to logarithmic phase in minimal medium. The enzyme was not present in the poorly transformable strain 23. The endonuclease was stable to 100 C for 30 min and, in a crude extract, was activated by heating at 80, 90, or 100 C. The activation caused a 5- to 10-fold increase in total units of enzyme activity. Sucrose gradient centrifugation indicated that the enzyme in a crude preparation has a major form (molecular weight, 66,000) which remains unchanged after heat activation. Under the assay conditions employed, the endonuclease did not release acid-soluble material from the substrate, high molecular weight tritiated deoxyribonucleic acid. The product, in double-stranded form, had a molecular weight of approximately 10(5), but it appeared to have undergone single-strand breaks.

Bacillus subtilis↗
PubMed1969

Gene-enzyme relationships in histidine biosynthesis in Bacillus subtilis.

Mutants for 9 of the 10 steps in histidine biosynthesis have been isolated and identified by enzyme assay. Each locus has been mapped in relation to the aro cluster and to other histidine loci by deoxyribonucleic acid-mediated transformation. The genes which code for enzymes 3, 6, and 8 of the pathway are linked to the aro cluster. A major histidine linkage group is composed of the genes which specify enzymes 1, 2, 5, 7, and 10. The locus which codes for step 9 of the pathway is unlinked to any other identified his loci. The major histidine cluster is loosely linked to cysB and is unlinked to any of the loci concerned with aromatic amino acid biosynthesis.

Bacillus subtilis↗
PubMed1969

Regulated enzymes of aromatic amino acid synthesis: control, isozymic nature, and aggregation in Bacillus subtilis and Bacillus licheniformis.

Several regulated enzymes involved in aromatic amino acid synthesis were studied in Bacillus subtilis and B. licheniformis with reference to organization and control mechanisms. B. subtilis has been previously shown (23) to have a single 3-deoxy-d-arabinoheptulosonate 7-phosphate (DAHP) synthetase but to have two isozymic forms of both chorismate mutase and shikimate kinase. Extracts of B. licheniformis chromatographed on diethylaminoethyl (DEAE) cellulose indicated a single DAHP synthetase and two isozymic forms of chorismate mutase, but only a single shikimate kinase activity. The evidence for isozymes has been supported by the inability to find strains mutant in these activities, although strains mutant for the other activities were readily obtained. DAHP synthetase, one of the isozymes of chorismate mutase, and one of the isozymes of shikimate kinase were found in a single complex in B. subtilis. No such complex could be detected in B. licheniformis. DAHP synthetase and shikimate kinase from B. subtilis were feedback-inhibited by chorismate and prephenate. DAHP synthetase from B. licheniformis was also feedback-inhibited by these two intermediates, but shikimate kinase was inhibited only by chorismate. When the cells were grown in limiting tyrosine, the DAHP synthetase, chorismate mutase, and shikimate kinase activities of B. subtilis were derepressed in parallel, but only DAHP synthetase and chorismate mutase were derepressible in B. licheniformis. Implications of the differences as well as the similarities between the control and the pattern of enzyme aggregation in the two related species of bacilli were discussed.

Bacillus↗
PubMed1968

Purification of competent cells in the Bacillus subtilis transformation system.

Transformed cells have been separated from nontransformed cells by centrifugation on a density gradient of Renografin-76. Separation was achieved both on a linear gradient and on a discontinuous gradient. Under optimal conditions, all of the cells in one band (median density, 1.110 g/ml) were transformants, whereas virtually all cells in the other (median density, 1.131) were nontransformants. In some instances, recentrifugation of the transformant band further enriched the transformant population. The transformed population can also be enriched by zonal centrifugation in a linear gradient of Ficoll. However, this technique is far less efficient than centrifugation in Renografin-76. Since the density of competent cells is identical to that of transformants, we conclude that the low density is a property of competent cells. The significance of this low density to the physiology of competent cells is discussed.

Bacillus subtilis↗
PubMed1968

Cross pathway regulation: effect of histidine on the synthesis and activity of enzymes of aromatic acid biosynthesis in Bacillus subtilis.

l-Histidine and, to a lesser degree, l-phenylalanine at concentrations of 10(-4)m inhibit the growth of leaky mutants (bradytrophs) of Bacillus subtilis that are deficient in the synthesis of p-hydroxyphenylpyruvate, the first intermediate specific to tyrosine synthesis. The inhibition can be overcome by growth factor amounts of l-tyrosine and p-hydroxyphenylpyruvate. Histidine and phenylalanine are capable of inhibiting the synthesis of tyrosine in several ways, and the major physiological effect which results in growth inhibition has not been established. Both l-histidine and l-phenylalanine inhibit the activity of prephenate dehydrogenase at concentrations about 100-fold higher than the inhibitory concentration of l-tyrosine. Histidine also appears to repress the synthesis of prephenate dehydrogenase because a histidine bradytroph growing in histidine-supplemented medium has a twofold lower level of this enzyme than the same cells growing in unsupplemented medium. These same two amino acids also inhibit the growth of a bradytroph deficient in dehydroquinate synthetase, an early enzyme in the pathway of tyrosine, phenylalanine, and tryptophan synthesis. The inhibition is overcome by a combination of tyrosine and phenylalanine. Histidine-resistant derivatives of both the prephenate dehydrogenase and dehydroquinate synthetase-deficient strains, which simultaneously have gained resistance to phenylalanine, have been isolated. Most of these resistant mutants synthesize additional tyrosine compared with the parent strain. One class of resistant mutants excretes tyrosine and has a number of enzymes of aromatic acid synthesis which are no longer repressible by any combination of the aromatic amino acids. Tyrosine inhibits the growth of histidine bradytrophs. Histidine, at growth factor levels, overcomes the inhibition.

Bacillus subtilis↗
PubMed1968

Common element in the repression control of enzymes of histidine and aromatic amino acid biosynthesis in Bacillus subtilus.

Single-step mutants of Bacillus subtilis derepressed for enzymes of both aromatic amino acid and histidine biosynthesis were isolated. These mutants occur at a frequency of 10(-6) per cell per generation. All histidine enzymes as well as all enzymes of aromatic acid synthesis which were examined are maximally derepressed. This level cannot be repressed by growth on either histidine or tyrosine. Some of the structural genes which specify the derepressed enzymes are linked to the aromatic cluster; others are unlinked. The significance of these nonrepressible strains is discussed in terms of the mechanism of repression.

Adenosine Triphosphate↗