RAPID ENZYME ASSAYS INVESTIGATING THE VARIATION IN THE GLYCOLYTIC PATHWAY IN FIELD-CAUGHT POPULATIONS OF FUNDULUS-HETEROCLITUS

Variation in enzyme expression may be important in evolutionary adaptation, yet is seldom studied. Furthermore, no studies have examined the expression of all enzymes in a defined metabolic pathway. Enzyme concentration is a measure of enzyme expression and was ascertained by assaying maximal activity. Presented here is an analysis of variation of maximal enzyme activity for all the enzymes in a single metabolic pathway, glycolysis, from three clinically distributed populations of the fish, Fundulus heteroclitus. Techniques for rapidly analyzing maximal enzyme activity for all the enzymes of an entire metabolic pathway from many individuals are described. The high degree of repeatability (mean coefficient of variation for replicates, 4.4%) and sensitivity (less than 3 mg of tissue is required to measure all 10 enzymes) of these assays demonstrate the utility of such an approach for analyzing variation among populations for a large numbers of enzymes. Results from these studies indicate that (1) the average coefficient of variation for all enzyme determinations within a population is 45.3% and (2) between populations, the activity of 5 of the 10 glycolytic enzymes are significantly different. This considerable variation occurs even in populations where there is little allelic variation. These data demonstrating; substantial variation in enzyme expression support the idea that changes in gene regulation may be as important as, or even more important than, changes in biochemical kinetic parameters in evolutionary processes.