L-Aspartase from Escherichia coli: substrate specificity and role of divalent metal ions

• Published in: Biochemistry (Easton) Vol. 27; no. 26; pp. 9089 - 9093
• Main Authors: Falzone, Christopher J, Karsten, William E, Conley, Judith D, Viola, Ronald E
• Format: Journal Article
• Language: English
• Published: Washington, DC American Chemical Society 27.12.1988
• Subjects: Ammonia-Lyases - metabolism; Analytical, structural and metabolic biochemistry; Aspartate Ammonia-Lyase - antagonists & inhibitors; Aspartate Ammonia-Lyase - metabolism; Aspartic Acid - analogs & derivatives; Aspartic Acid - metabolism; Bacterial Proteins - antagonists & inhibitors; Bacterial Proteins - metabolism; Binding Sites; Biological and medical sciences; Cations, Divalent - metabolism; Electron Spin Resonance Spectroscopy; Enzyme Activation - drug effects; Enzymes and enzyme inhibitors; Escherichia coli - enzymology; Fundamental and applied biological sciences. Psychology; Hydrogen-Ion Concentration; Protein Binding; Substrate Specificity
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/bi00426a004

The enzyme L-aspartase from Escherichia coli has an absolute specificity for its amino acid substrate. An examination of a wide range of structural analogues of L-aspartic acid did not uncover any alternate substrates for this enzyme. A large number of competitive inhibitors of the enzyme have been characterized, with inhibition constants ranging over 2 orders of magnitude. A divalent metal ion is required for enzyme activity above pH 7, and this requirement is met by many transition and alkali earth metals. The binding stoichiometry has been established to be one metal ion bound per subunit. Paramagnetic relaxation studies have shown that the divalent metal ion binds at the recently discovered activator site on L-aspartase and not at the enzyme active site. Enzyme activators are bound within 5 A of the enzyme-bound divalent metal ion. The activator site is remote from the active site of the enzyme, since the relaxation of inhibitors that bind at the active site is not affected by paramagnetic metal ions bound at the activator site.