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Z Barak

Publications and source records attributed to Z Barak.

52 records · Page 3Linked to original sources
PubMed1987

A method for simultaneous determination of the two possible products of acetohydroxy acid synthase.

A method for the simultaneous assay of 2-acetolactate and 2-aceto-2-hydroxybutyrate formation catalyzed by acetohydroxy acid synthase in the presence of its substrates pyruvate and 2-ketobutyrate is described. The method, appropriate for the study of the physiologically and mechanistically significant competition between the two reactions, involves oxidative decarboxylation of the acetohydroxy acids to the corresponding 2,3-diketones, transfer of the volatile diketones to methanol, and gas chromatographic analysis with electron-capture detection. Oxidative decarboxylation by air requires catalytic activation, and addition of iron salts is crucial to the success of the method with purified enzymes.

Acetolactate Synthase↗
PubMed1987

Fate of Bacillus thuringiensis subsp. israelensis under Simulated Field Conditions.

The fate of Bacillus thuringiensis subsp. israelensis in a natural aquatic habitat was studied in a model system by using laboratory-simulated field waters and a mutant of the bacterium resistant to three antibiotics. Contact with mud of a sporal culture of the mutant resulted in an immediate disappearance of the larvicidal activity but had no influence on viability. The cessation of toxicity was caused by bacterial adsorption on soil particles, since 99.8% of the bacteria was found in the mud fraction within 45 min, with concurrent disappearance from the supernatant. When the mud was stirred, the bacteria could be redetected. The viability count of the mud suspension remained practically constant for at least 22 days, indicating that the spores were still fully viable but were incapable of germinating and multiplying in the mud under our experimental conditions. Approximately 8% of the colony forming ability of the bacteria could be separated from the mud by vigorous mixing followed by immediate filtration. The filtrated spores retained their toxicity, killing 90% of the larval populations even after 22 days incubation in the soil. The inactivation of the toxic activity of B. thuringiensis subsp. israelensis in the mud was therefore a reversible process and was probably due to masking of the bacteria, thus making the bacteria and their toxin inaccessible to the larvae. In the simulated field waters without mud, we observed only a very slow inhibition of the larvicidal activity. In contrast to the activity in the mud suspension, this activity could not be restored.

Journal Article↗
PubMed1987

Physiological implications of the specificity of acetohydroxy acid synthase isozymes of enteric bacteria.

The rates of formation of the two alternative products of acetohydroxy acid synthase (AHAS) have been determined by a new analytical method (N. Gollop, Z. Barak, and D. M. Chipman, Anal. Biochem., 160:323-331, 1987). For each of the three distinct isozymes of AHAS in Escherichia coli and Salmonella typhimurium, a specificity ratio, R, was defined: Formula: see text, which is constant over a wide range of substrate concentrations. This is consistent with competition between pyruvate and 2-ketobutyrate for an active acetaldehyde intermediate formed irreversibly after addition of the first pyruvate moiety to the enzyme. Isozyme I showed no product preference (R = 1), whereas isozymes II and III form acetohydroxybutyrate (AHB) at approximately 180- and 60-fold faster rates, respectively, than acetolactate (AL) at equal pyruvate and 2-ketobutyrate concentrations. R values higher than 60 represent remarkably high specificity in favor of the substrate with one extra methylene group. In exponentially growing E. coli cells (under aerobic growth on glucose), which contain about 300 microM pyruvate and only 3 microM 2-ketobutyrate, AHAS I would produce almost entirely AL and only 1 to 2% AHB. However, isozymes II and III would synthesize AHB (on the pathway to Ile) and AL (on the pathway to valine-leucine) in essentially the ratio required for protein synthesis. The specificity ratio R of any AHAS isozyme was affected neither by the natural feedback inhibitors (Val, Ile) nor by the pH. On the basis of the specificities of the isozymes, the known regulation of AHAS I expression by the catabolite repression system, and the reported behavior of bacterial mutants containing single AHAS isozymes, we suggest that AHAS I enables a bacterium to cope with poor carbon sources, which lead to low endogenous pyruvate concentrations. Although AHAS II and III are well suited to producing the branched-chain amino acid precursors during growth on glucose, they would fail to provide appropriate quantities of AL when the concentration of pyruvate is relatively low.

Acetolactate Synthase↗
PubMed1986

Biological control of mosquitoes by the larvicidal activity of Bacillus thuringiensis var. israelensis delta endotoxin.

Bacillus thuringiensis var. israelensis (B.t.i.) is a promising, safe toxic agent for control of mosquitoes, but the rapid disappearance of its toxicity makes its use in practice economically unattractive. The lack of evidence for B.t.i. multiplication in water also makes the natural ecology of B.t.i. puzzling. The observation that mosquito larvae readily cannibalize carcasses of B.t.i.-killed larvae, and that the carcasses become toxic to scavenging larvae provides a possible solution to this puzzle. Several experimental techniques have been developed to study these phenomena (e.g., quantitative determination of spore numbers despite aggregation, protocols for following toxicity development, etc.). Results suggest a cycle involving larval poisoning and death due to delta-endotoxin of ca. 1,000 spores, germination, vegetative growth and sporulation (with toxin production) of B.t.i. after multiplication to several million in the carcass. Implications of these results for the ecology of B.t.i., and for practical applications in mosquito control, are discussed.

Animals↗
PubMed1983

Inhibition of acetohydroxy acid synthase by leucine.

The enzymatic reaction of acetohydroxy acid synthase in crude extracts of Escherichia coli K-12 is inhibited by leucine. Inhibition is most pronounced at low pH values and is low at pH values higher than 8.0. Both isoenzymes of acetohydroxy acid synthase present in E. coli K-12 (isoenzyme I and isoenzyme III) are inhibited by leucine. Isoenzyme I, which is responsible for the majority of acetohydroxy acid synthase activity in E. coli K-12 at physiological pH, is inhibited almost completely by 30 mM leucine at pH 6.25-7.0 and is not affected at all at pH values higher than 8.4. Inhibition of isoenzyme I by leucine is a mixed noncompetitive process. Leucine inhibition of isoenzyme III is pH-independent and reaches only 40% at 30 mM leucine. The inhibition of acetohydroxy acid synthase by leucine at physiological pH, observed in vitro in this study, correlates with the idea that acetohydroxy acid synthase is a target for the toxicity of the abnormally high concentrations of leucine in E. coli K-12.

Acetolactate Synthase↗
PubMed1981

Toxicity of leucine-containing peptides in Escherichia coli caused by circumvention of leucine transport regulation.

A variety of leucine-containing peptides (LCP), Phe-Leu, Gly-Leu, Pro-Leu, Ala-Leu, Ala-Leu-Lys, Leu-Phe-Ala, Leu-Leu-Leu, and Leu-Gly-Gly, inhibited the growth of a prototrophic strain of Escherichia coli K-12 at concentrations between 0.05 and 0.28 mM. Toxicity requires normal uptake of peptides. When peptide transport was impaired by mutations, strains became resistant to the respective LCP. Inhibition of growth occurred immediately after the addition of LCP, and was relieved when 0.4 mM isoleucine was added. The presence of Gly-Leu in the medium correlated with the inhibition of growth, and the bacteria began to grow at the normal rate 70 min after Gly-Leu became undetectable. Disappearance of the peptide corresponded with the appearance of free leucine and glycine in the medium. The concentration of leucine inside the LCP-treated bacteria was higher than that in the leucine-treated and the control cultures. We suggest that entry of LCP into the cells via peptide transport systems circumvents the regulation of leucine transport, thereby causing abnormality high concentrations of leucine inside the cells. This accumulation of leucine interferes with the biosynthesis of isoleucine and inhibits the growth of the bacteria.

Biological Transport↗
PubMed1975

Peptide transport.

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Amino Acid Sequence↗
PubMed1975

Specialized peptide transport system in Escherichia coli.

Trileucine is utilized as a source of leucine for growth of strains of Escherichia coli K-12 that are deficient in the oligopeptide transport system (Opp). Trithreonine is toxic to E. coli K-12. Opp- mutants of E. coli K-12 retain complete sensitivity to this tripeptide. Moreover, E. coli W, which is resistant to trithreonine, can utlize this tripeptide as a threonine source and this capability is fully maintained in E. coli W (Opp-). A spontaneous trithreonine-resistant mutant of E. coli K-12 (Opp-) has been isolated that has an impaired growth response to trileucine and is resistant to trithreonine. Trileucine competes with the uptake of trithreonine as measured by its ability to relieve trithreonine toxicity in E. coli K-12. It is concluded that trileucine as well as trithreonine are transported into E. coli K-12 or W by a common uptake system that is distinct from the Opp system. Trimethionine can act as a competitor of trileucine or trithreonine-supported growth and as an antagonist of trithreonine toxicity in Opp- mutants. It is concluded that trimethionine is recognized by the trileucine-trithreonine transport system. Trithreonine, trimethionine, and trileucine are also transported by the Opp system, as they all relieve triornithine toxicity towards E. coli W and compete with tetralysine utilization as lysine source for growth of a lysine auxotroph of this strain.

Biological Transport, Active↗