Humic acids-tyrosinase interactions as a model of soil humic-enzyme complexes

Abstract

Interactions between tyrosinase and humic acids (HA), which are of great importance for soil biochemistry, have been studied with respect to binding mechanisms involved and to alterations of the activity and of some properties of the enzyme. Neutralized solutions of soil HA slightly inhibited tyrosinase activity. The extent of inhibition (up to 14.0%) was related to HA concentration. Stable and insoluble HA-tyrosinase complexes were obtained by coflocculation of HA and enzyme with Ca 2+. Depending on the HA concentration and probably on the pH, from 32 to 76% of the added enzyme was strongly bound. The data from i.r. spectroscopy suggest the involvement of carboxyl and phenolic groups of HA in ionic and hydrogen bond formation with the enzyme molecule. Some properties of the HA-tyrosinase complexes and the soluble enzyme were also compared. When tyrosinase was bound by HA, it showed a substrate specificity similar to that of the free enzyme but a K m value slightly higher and a thermal stability and resistance to proteolysis lower than those observed for the free enzyme. Although the systems studied were artificial, they are helpful in understanding the nature of humic-enzyme complexes which apparently occur in the soil.