Abstract: This chapter describes the assay, purification, and properties of glutamine synthetase. The synthesis of glutamine in microorganisms may be regarded as the first step in a highly branched pathway, which leads ultimately to the biosynthesis of a large number of different compounds, including tryptophan, adenylic acid, cytidylic acid, and glucosamine 6-phosphate. Glutamine synthetase from Escherichia coli is, therefore, of cardinal importance in biosynthetic metabolism and is a strategic target for cellular regulation. The purified enzyme may be assayed by measuring the production of inorganic phosphate in the biosynthetic reaction. Alternatively, a continuous recording of catalytic activity may be achieved by coupling the production of adenosine diphosphate (ADP) in the biosynthetic assay to the oxidation of diphosphopyridine nucleotide (DPNH) by the addition of phosphoenolpyruvate, pyruvate kinase, and lactate dehydrogenase in excess. Preparations of purified glutamine synthetase may differ from one another in the amount of covalently bound adenosine monophosphate (AMP) residues that they contain. Mn2+ stabilizes the native dodecameric structure of the enzyme thus, protecting the enzyme from reacting with sulfhydryl reagents and other mild denaturants.
Publication Year: 1970
Publication Date: 1970-01-01
Language: en
Type: book-chapter
Indexed In: ['crossref']
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Cited By Count: 452
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