RATIONAL ENGINEERING OF ACTIVITY AND SPECIFICITY IN A SERINE-PROTEASE

The discovery of the Na+-dependent allosteric regulation in serine pro teases makes it possible to control catalytic activity and specificity in this class of enzymes in a way never considered before. We demonst rate that rational site-directed mutagenesis of residues controlling N a+ binding can profoundly alter the properties of a serine protease. B y suppressing Na+ binding to thrombin, we shift the balance between pr ocoagulant and anticoagulant activities of the enzyme. Those mutants, compared to wild-type, have reduced specificity toward fibrinogen, but enhanced or slightly reduced specificity toward protein C. Because th is engineering strategy targets a fundamental regulatory mechanism, it is amenable of extension to other enzymes of biological and pharmacol ogical importance.