[Production and catalytic properties of point mutants Phe41--->His and Phe143--->Glu of horseradish peroxidase, expressed in Escherichia coli].

Recombinant horseradish peroxidase and its single-point mutants, F4IH and H143E, have been reactivated from E. coli inclusion bodies with a 25% yield. Both mutations affect heme entrapment as well as enzyme stability and activity. A more than 40-fold decrease in the specific activity towards ABTS is associated with different steps of peroxidase catalysis. F41H replacement results in a drop of both rate constants by two and one orders or magnitude for hydrogen peroxide and ABTS, respectively. The mechanism of iodide oxidation by the F41H mutant fits into a ternary interaction. The F143E replacement mainly affects the steps of ABTS oxidation and product dissociation. It is suggested that the role of replaced phenylalanine residues consists in the formation of a highly hydrophobic pocket allowing for strong non-covalent binding of the heme porphyrin ring.