Crystal structure of bovine lipoyltransferase in complex with lipoyl-AMP.

Kazuko Fujiwara, Harumi Hosaka, Makoto Matsuda, Kazuko Okamura-Ikeda, Yutaro Motokawa, Mamoru Suzuki, Atsushi Nakagawa, Hisaaki Taniguchi

Institute for Enzyme Research, the University of Tokushima, Tokushima 770-8503, Japan. fujiwara@ier.tokushima-u.ac.jp

Journal of molecular biology 2007 Aug 3

filter terms:

Lipoic acid is an essential cofactor of the alpha-ketoacid dehydrogenase complexes and the glycine cleavage system. It is covalently attached to a specific lysine residue of the subunit of the complexes. The bovine lipoyltransferase (bLT) catalyzes the lipoic acid attachment reaction using lipoyl-AMP as a substrate, forming a lipoylated protein and AMP. To gain insights into the reaction mechanism at the atomic level, we have determined the crystal structure of bLT at 2.10 A resolution. Unexpectedly, the purified recombinant bLT contains endogenous lipoyl-AMP. The structure of bLT consists of N-terminal and C-terminal domains, and lipoyl-AMP is bound to the active site in the N-terminal domain, adopting a U-shaped conformation. The lipoyl moiety is buried in the hydrophobic pocket, forming van der Waals interactions, and the AMP moiety forms numerous hydrogen bonds with bLT in another tunnel-like cavity. These interactions work together to expose the C10 atom of lipoyl-AMP to the surface of the bLT molecule. The carbonyl oxygen atom of lipoyl-AMP interacts with the invariant Lys135. The interaction might stimulate the positive charge of the C10 atom of lipoyl-AMP, and consequently facilitate the nucleophilic attack by the lysine residue of the lipoate-acceptor protein, accompanying the bond cleavage between the carbonyl group and the phosphate group. We discuss the structural differences between bLT and the lipoate-protein ligase A from Escherichia coli and Thermoplasma acidophilum. We further demonstrate that bLT in mitochondria also contains endogenous lipoylmononucleotide, being ready for the lipoylation of apoproteins.

Citation

Kazuko Fujiwara, Harumi Hosaka, Makoto Matsuda, Kazuko Okamura-Ikeda, Yutaro Motokawa, Mamoru Suzuki, Atsushi Nakagawa, Hisaaki Taniguchi. Crystal structure of bovine lipoyltransferase in complex with lipoyl-AMP. Journal of molecular biology. 2007 Aug 3;371(1):222-34