Effect of chloride and glutamate ions on in vitro protein synthesis by the moderate halophile Vibrio costicola

Abstract

Vibrio costicola grown in the presence of different NaCl concentrations contains cell-associated Na+ and K+ ions whose sum is equal to or greater than the external Na+ concentration. In the presence of 0.5 M NaCI, virtually no in vitro protein is synthesized in extracts of cells grown in 1.0 M NaCI. However, we report here that active in vitro protein synthesis occurred in 0.6 M or higher concentrations of Na2SO4, sodium formate, sodium acetate, sodium aspartate, or sodium glutamate, whereas 0.6 M NaF, NaCl, or NaBr completely inhibited protein synthesis as measured by polyuridylic acid-directed incorporation of ['4C]phenylalanine. Sodium glutamate, sodium aspartate, and betaine (0.3 M) counteracted the inhibitory action of 0.6 M NaCl. The cell-associated Cl-concentration was 0.22 mol/kg in cells grown in 1.0 M NaCl. Of this, the free intracellular Cl-concentration was only 0.02 mol/kg. Cells contained 0.11 mol of glutamate per kg and small concentrations of other amino acids. All of the negative counterions for cell-associated Na+ and K+ have not yet been determined. In vitro protein synthesis by Escherichia coli was inhibited by sodium glutamate. Hybridization experiments with ribosomes and the soluble (S-100) fractions from extracts of E. coli and V. costicola showed that the glutamate-sensitive fraction was found in the soluble, not the ribosomal, part of the system. The phenylalanyl-tRNA synthetase of V. costicola was not inhibited by 0.5 M or higher concentrations of NaCI; it was slightly more sensitive to high concentrations of sodium glutamate. Therefore, this enzyme was not responsible for the salt response of the V. costicola in vitro protein-synthesizing system.

Low-Concentration Kinetics of Atmospheric CH 4 Oxidation in Soil and Mechanism of NH 4 + Inhibition

“…All of the ions used in this study potentially could affect CH 4 oxidation in three basic ways. First, they could change the soil osmotic potential and impose water stress on the microbial community (18,40); second, they could affect ion exchange, thereby altering NH 4 ϩ availability (28); and third, they could affect the CH 4 oxidizers directly in a number of ways (11,15,21). Any of these factors could alter CH 4 oxidation rates and kinetics and thus affect the interpretation of the specific NH 4 ϩ inhibition mechanism.…”

Molecular adaptation of enzymes, metabolic systems and transport systems in halophilic bacteria

“…Direct measurements showed that levels of CI-ions were much lower than K + or Na + ions in cells of these organism. If, instead of NaC1 or KC1, Na-or K-glutamate was used, maximal activity occurred in 0.6 M salt, and substantial activity remained at 0.9 M [16].…”

“…It appears from these results that C1-ions inhibit protein synthesis. It should be noted that these ions are excluded from E. coil, yeast and animal cells (reviewed in [16]). They are not excluded from the halobacteria, however, being found in as high concentrations inside as outside the cells [5,6].…”

“…The site of CI-action is still unknown. It has been shown not to act on phenylalanyl tRNA synthetase [16]; there is some preliminary evidence that it prevents binding of mRNA to ribosomes. Other possible sites of action have not yet been explored.…”

“…Effects of salts on in vitro protein synthesis by V. costicolaData from[16]. 100% activity -~ 31.9 pmol of [14C]phenylalanine (32500 clam) incorporated into hot tfichloroacetic acid-insoluble material.…”