Destruction and reformation of an iron-sulfur cluster during catalysis by lipoyl synthase.

Abstract

Lipoyl synthase (LipA) catalyzes the last step in the biosynthesis of the lipoyl cofactor, which is the attachment of two sulfhydryl groups to C6 and C8 of a pendant octanoyl chain. The appended sulfur atoms derive from an auxiliary [4Fe-4S] cluster on the protein that is degraded during turnover, limiting LipA to one turnover in vitro. We found that the <i>Escherichia coli</i> iron-sulfur (Fe-S) cluster carrier protein NfuA efficiently reconstitutes the auxiliary cluster during LipA catalysis in a step that is not rate-limiting. We also found evidence for a second pathway for cluster regeneration involving the <i>E. coli</i> protein IscU. These results show that enzymes that degrade their Fe-S clusters as a sulfur source can nonetheless act catalytically. Our results also explain why patients with <i>NFU1</i> gene deletions exhibit phenotypes that are indicative of lipoyl cofactor deficiencies.