Reactivity of manganese peroxidase: Site-directed mutagenesis of residues in proximity to the porphyrin ring

The purpose of this study was to determine the effect of heme pocket hydrop hobicity on the reactivity of manganese peroxidase, Residues within 5 Angst rom of the heme active site were identified. From this group, Leu169 and Se r172 were selected and mutated to Phe and Ale, respectively. The mutant pro teins were then characterized by steady-state kinetics. Whereas the Leu169P he mutation had little, if any, effect on activity, the Ser172Ala mutation decreased k(cat) and also the specificity constant (k(cat)/K-m) for Mn2+, b ut not H2O2. Transient-state studies indicated that the mutation affected o nly the reactions of compound II. These results indicate that compound II i s the most sensitive to changes in the heme environment. (C) 2000 Academic Press.