The Study of Enzyme Reactions at Subzero Temperatures

Abstract

While there is a considerable volume of direct evidence that sequential enzyme-substrate complexes are involved in enzyme-catalyzed reactions in general, structural information about such transient and very fleeting complexes is scarce. Fast kinetic techniques now permit us to carry out measurements in a very short interval of time but most of these techniques give nothing more than characteristic relaxation times with no indication about what these times refer to in terms of stereochemical changes of the reacting system. Since it is obvious that conformational changes are required for the enzyme action lack of information about such changes at the molecular level does not permit us to go further in the understanding of reaction mechanisms. A detailed structure analysis of the intermediate complexes could lead to a direct observation of enzyme processes at the molecular level, after a suitable “quenching” of such processes at given intermediate steps. Since most enzyme reactions proceed in steps with energy barriers which are the primary determinants of how rapidly these reactions occur at biological temperatures, there is great potential interest in the use of subzero temperatures to stabilize intermediate enzyme-substrate complexes. The rate at which reactions occur is largely set by the free energy of activation and this energy can be regarded as the amount of energy which must be put into each enzyme-substrate complex to be stabilized and the reaction quenched.