Properties of the Bacillus licheniformis A5 glutamine synthetase purified from cells grown in the presence of ammonia or nitrate

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Abstract
The glutamine synthetase from B. licheniformis A5 was purified by using a combination of polyethylene glycol precipitation and chromatography on Bio-Gel A 1.5m. The resulting preparation was judged to be homogeneous by the criteria of polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, equilibrium analytical ultracentrifugation and EM analysis. The enzyme is a dodecamer with a MW of .apprx. 16,000 and its subunit MW is 51,000. Under optimal assay conditions (pH 6.6, 37.degree. C) apparent Km values for glutamate, ammonia and MnATP (1:1 ratio) were 3.6, 0.4 and 0.9 mM, respectively. Glutamine synthetase activity was inhibited .apprx. 50% by the addition of 5 mM glutamine, alanine, glycine, serine, .alpha.-ketoglutarate, carbamyl phosphate, ADP or ITP to the standard glutamine synthetase assay system; 5 mM AMP or pyrophosphate caused .apprx. 90% inhibition of enzyme activity. Phosphorylribosyl pyrophosphate at 5 mM enhanced activity .apprx. 60%. No physical or kinetic differences in the properties of the enzyme when it was purified from cells grown in the presence of ammonia or nitrate as sole N source were detected. B. licheniformis A5 apparently contains 1 species of glutamine synthetase whose catalytic activity is not regulated by a covalent modification system.

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