Solution structure of the lipoyl domain of the chimeric dihydrolipoyl dehydrogenase P64K from Neisseria meningitidis

The antigenic P64K protein from the pathogenic bacterium Neisseria meningit idis is found in the outer membrane of the cell, and consists of two parts: an 81-residue N-terminal region and a 482-residue C-terminal region. The a mino-acid sequence of the N-terminal region is homologous with the lipoyl d omains of the dihydrolipoyl acyltransferase (E2) components, and that of th e C-terminal region with the dihydrolipoyl dehydrogenase (E3) components, o f 2-oxo acid dehydrogenase multienzyme complexes. The two parts are separat ed by a long linker region, similar to the linker regions in the E2 chains of 2-oxo acid dehydrogenase complexes, and it is likely this region is conf ormationally flexible. A subgene encoding the P64K lipoyl domain was create d and over-expressed in Escherichia coli. The product was capable of post-t ranslational modification by the lipoate protein ligase but not aberrant mo dification by the biotin protein ligase of E. coli. The solution structure of the apo-domain was determined by means of heteronuclear NMR spectroscopy and found to be a flattened beta barrel composed of two four-stranded anti parallel beta sheets. The lysine residue that becomes lipoylated is in an e xposed beta turn that, from a {H-1}-N-15 heteronuclear Overhauser effect ex periment, appears to enjoy substantial local motion. This structure of a li poyl domain derived from a dihydrolipoyl dehydrogenase resembles that of li poyl domains normally found as part of the dihydrolipoyl acyltransferase co mponent of 2-oxo acid dehydrogenase complexes and will assist in furthering the understanding of its function in a multienzyme complex and in the memb rane-bound P64K protein itself.