Structures of lipoyl synthase reveal a compact active site for controlling sequential sulfur insertion reactions

DOI: 10.1042/BJ20140895
PMID: 25100160

Authors: Jenny Harmer (University of Southampton) , Martyn Hiscox (University of Southampton) , Pedro Dinis (University of Southampton) , Stephen Fox (University of Southampton) , Andreas Iliopoulos (University of Southampton) , James Hussey (University of Southampton) , James Sandy (Diamond Light Source) , Florian Van Beek (University of Southampton) , Jonathan Essex (University of Southampton) , Peter Roach (University of Southampton)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Biochemical Journal

State: Published (Approved)
Published: August 2014

Abstract: Lipoyl cofactors are essential for living organisms and are produced by the insertion of two sulfur atoms into the relatively unreactive C-H bonds of an octanoyl substrate. This reaction requires lipoyl synthase, a member of the radical SAM enzyme superfamily. Herein we present crystal structures of lipoyl synthase with two [4Fe-4S] clusters bound at opposite ends of the TIM barrel, the usual fold of the radical SAM superfamily. The cluster required for reductive SAM cleavage conserves the features of the radical SAM superfamily, but the auxiliary cluster is bound by a CX4CX5C motif unique to lipoyl synthase. The fourth ligand to the auxiliary cluster is an extremely unusual serine residue. Site directed mutants show this conserved serine ligand is essential for the sulfur insertion steps. One crystallized LipA complex contains MTA, a breakdown product of SAM, bound in the likely SAM binding site. Modelling has identified an 18 Å deep channel, well-proportioned to accommodate an octanoyl substrate. These results suggest the auxiliary cluster is the likely sulfur donor, but access to a sulfide ion for the second sulfur insertion reaction requires the loss of an iron atom from the auxiliary cluster, which the serine ligand may enabled.

Journal Keywords: cofactor; crystal structure; enzyme catalysis; lipoyl synthase; radical S-adenosylmethionine (SAM); sulfur; Thermosynechococcus elongatus

Diamond Keywords: Enzymes; Cyanobacteria; Bacteria

Subject Areas: Biology and Bio-materials, Chemistry


Instruments: I02-Macromolecular Crystallography

Added On: 12/08/2014 14:20

Discipline Tags:

Biochemistry Catalysis Chemistry Structural biology Life Sciences & Biotech

Technical Tags:

Diffraction Macromolecular Crystallography (MX)