EFFECT OF A NEGATIVE CHARGE ON THE SCREENING OF THE ACTIVE-SITE OF HORSERADISH-PEROXIDASE

The F143E mutant form of the recombinant horseradish peroxidase was re activated from E. coli inclusion bodies. The mutation inhibits heme en trapment and results in a decrease in the catalytic activity, mainly a ffecting the stage of the oxidation of a donor substrate (ABTS, iodide ). An increase in stability of the mutant form obtained under radiatio n inactivation over that of the wild-type recombinant enzyme was obser ved. The data obtained confirms the proposed location of Phe143 at the entrance of the active center, hence its replacement by the negativel y charged glutamic acid residue retards heme entrapment and substrate binding, thus protecting the active center of the enzyme against the r adicals generated by radiolysis.