Substrate activation and thermal denaturation kinetics of the tetrameric and the trypsin-generated monomeric forms of horse serum butyrylcholinesterase

Abstract

Native horse serum butyrylcholinesterase (acylcholine acylhydrolase; EC 3.1.1.8) is a tetrameric enzyme which can dissociate after a limited proteolysis by trypsin into three additional molecular forms, including the monomeric entity. The trypsin-generated monomer of butyrylcholinesterase, isolated by ultracentrifugation on sucrose gradient, is stable and allows the relations between the polymeric structure of butyrylcholinesterase and its kinetic characteristics to be approached, e.g., substrate activation and complex thermal denaturation curves. The trypsin-generated monomer of butyrylcholinesterase behaves with identical kinetic parameter values as the native tetrameric enzyme. On the other hand, the thermal denaturation of the native tetrameric butyrylcholinesterase does not follow first-order kinetics, but may be described by a sum of exponential terms. This behavior is not due to the polymeric nature of butyrylcholinesterase but seems to be related to a structural heterogeneity induced by the heat treatment.