title: A novel amidotransferase required for lipoic acid cofactor assembly in Bacillus subtilis.
abstract: In the companion paper we reported that Bacillus subtilis requires three proteins for lipoic acid metabolism, all of which are members of the lipoate protein ligase family. Two of the proteins, LipM and LplJ, have been shown to be an octanoyltransferase and a lipoate : protein ligase respectively. The third protein, LipL, is essential for lipoic acid synthesis, but had no detectable octanoyltransferase or ligase activity either in vitro or in vivo. We report that LipM specifically modifies the glycine cleavage system protein, GcvH, and therefore another mechanism must exist for modification of other lipoic acid requiring enzymes (e.g. pyruvate dehydrogenase). We show that this function is provided by LipL, which catalyses the amidotransfer (transamidation) of the octanoyl moiety from octanoyl-GcvH to the E2 subunit of pyruvate dehydrogenase. LipL activity was demonstrated in vitro with purified components and proceeds via a thioester-linked acyl-enzyme intermediate. As predicted, ΔgcvH strains are lipoate auxotrophs. LipL represents a new enzyme activity. It is a GcvH:[lipoyl domain] amidotransferase that probably uses a Cys-Lys catalytic dyad. Although the active site cysteine residues of LipL and LipB are located in different positions within the polypeptide chains, alignment of their structures show these residues occupy similar positions. Thus, these two homologous enzymes have convergent architectures.
authors.affiliation: Departments of Microbiology Biochemistry Chemistry Biology Interface Training Program, University of Illinois, Urbana, IL 61801, USA.
authors.forename: Quin H
authors.initials: QH
authors.lastname: Christensen
authors.forename: Natalia
authors.initials: N
authors.lastname: Martin
authors.forename: Maria C
authors.initials: MC
authors.lastname: Mansilla
authors.forename: Diego
authors.initials: D
authors.lastname: de Mendoza
authors.forename: John E
authors.initials: JE
authors.lastname: Cronan
mesh_terms: D001412:Bacillus subtilis / Q000201:enzymology* / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D053898:Biosynthetic Pathways / Q000235:genetics*; D017353:Gene Deletion; D005798:Genes, Bacterial; D011768:Pyruvate Dehydrogenase Complex / Q000378:metabolism; D008063:Thioctic Acid / Q000096:biosynthesis*
chemical_list: D001426:Bacterial Proteins; D011768:Pyruvate Dehydrogenase Complex; D008063:Thioctic Acid