PMID:8875647
| Citation |
Drueckes, P and Schinzel, R (1996) Activation of E350A mutant maltodextrin phosphorylase by exogenously added acetate. Protein Eng. 9:701-5 |
|---|---|
| Abstract |
Site-directed mutagenesis of E350 to alanine in Escherichia coli maltodextrin phosphorylase reduced both enzyme activity (100-fold) and apparent binding of the oligosaccharide substrate (10-fold), suggesting a participation of this residue in binding of the substrate in the ground and transition states. The E350A mutant enzyme was found to be activated up to 20-fold by exogenous acetate ions which substitute for the deleted side chain. In contrast, apparent binding was not affected by acetate ions, indicating a dual role for the carboxylic group of this residue in catalysis and binding. Formate also appears to activate the E350A mutant enzyme, but this effect is obscured by the strong inhibitory effect of formate on the wild-type enzyme. For propionate ions, a weak 2-fold activation was noticed, while other compounds like trifluoroacetate and acetamide had no effects on the catalytic properties of either the E350A mutant enzyme or wild-type enzyme. If E350 was substituted by a glutamine, no activation was observed upon the addition of acetate ions. However, a weak activation by formate was found, confirming that activation by acetate is caused by specific binding at the mutated site. |
| Links | |
| Keywords |
Acetic Acid/metabolism; Carboxylic Acids/pharmacology; Cations/pharmacology; Enzyme Activation; Escherichia coli; Glucosyltransferases/chemistry; Glucosyltransferases/metabolism; Mutagenesis, Site-Directed; Structure-Activity Relationship |
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