Abstract
Cysteine dioxygenase (CDO) is a non-heme monoiron enzyme with an unusual posttranslational modification in the proximity of the ferrous iron active site. This modification, a cysteine to tyrosine thioether bond, cross-links two beta-strands of the beta-barrel. We have investigated its role in catalysis through a combined crystallographic and kinetic approach. The C93G variant lacks the cross-link and shows little change in structure from that of the wild type, suggesting that the cross-link does not stabilize an otherwise unfavorable conformation. A pH-dependent kinetic study shows that both cross-linked and un-cross-linked CDO are active but the optimal pH decreases with the presence of the cross-link. This result reflects the effect of the thioether bond on the pK(a) of Y157 and this residues role in catalysis. At higher pH values, k(cat) is also higher for the cross-linked form, extending the pH range of activity. We therefore propose that the cross-link also increases activity by controlling deleterious interactions involving the thiol/ate of C93.