The Cofactor of Tetrachloroethene Reductive Dehalogenase of Dehalospirillum multivorans Is Norpseudo‐B₁₂, a New Type of a Natural Corrinoid

Abstract

Dedicated to Professor Duilio Arigoni on the occasion of his 75th birthdayThe corrinoid cofactor of the tetrachloroethene reductive dehalogenase of Dehalospirillum multivorans was isolated in its Cob-cyano form. This cofactor represents the main corrinoid found in D. multivorans cells. Analysis of the isolated cyano-corrinoid by a combination of HPLC and UV/VIS-absorbance spectroscopy revealed it to be nonidentical to a variety of known natural B 12 derivatives. From high-resolution massspectrometric analysis, the molecular formula of the corrinoid isolated from D. multivorans could be deduced as C 58 H 81 CoN 17 O 14 P. The sample of the novel corrinoid from D. multivorans was further analyzed by UV/VIS, CD, and one-and two-dimensional 1 H-, 13 C-, and 15 N-NMR spectroscopy, which indicated its structure to be closely related to that of pseudovitamin B 12 (Cob-cyano-7''-adeninylcobamide). By the same means, the corrinoid could be shown to differ from pseudovitamin B 12 only by the lack of the methyl group attached to carbon 176, and, therefore, it was named norpseudovitamin B 12 (or, more precisely, 176-norpseudovitamin B 12 ). Norpseudovitamin B 12 represents the first example of a −complete× B 12 -cofactor that lacks one of the methyl groups of the cobamide moiety, indicating that the B 12 -biosynthetic pathway in D. multivorans differs from that of other organisms. X-Ray crystal-structures were determined for norpseudovitamin B 12 from D. multivorans and the analogues pseudovitamin B 12 and factor A (Cob-cyano-7''-[2-methyl]adeninylcobamide). These first accurate crystal structures of complete corrinoids with an adeninyl pseudonucleotide confirmed the expected coordination properties around Co and corroborated the close conformational similarity of the nucleotide moieties of norpseudovitamin B 12 and its two homologues.

Retentive Memory of Bacteria: Long-Term Regulation of Dehalorespiration in Sulfurospirillum multivorans

“…This result was coincident with previous observations that in cells grown in the absence of PCE a significant portion of the enzyme is localized in the cytoplasm or at the cytoplasmic face of the cell membrane (9). Long-term subcultured cells, which had lost PceA, also lacked the norpseudovitamin B 12 cofactor as detected by corrinoid extraction (data not shown) (12).…”

“…The key enzyme of PCE utilization that converts PCE via TCE to cis-1,2-dichloroethene (cDCE) is the PCE reductive dehalogenase (PceA). Mature PceA is a monomeric enzyme harboring one norpseudovitamin B 12 and two Fe/S clusters as cofactors (12,15). The cytoplasmic precursor of the PCE dehalogenase (pre-PceA) bears an N-terminal signal peptide of 37 amino acids, including the motif SRRXFXK, which identifies pre-PceA as a substrate of the Tat (twin arginine translocation) protein export pathway (2).…”

Diversity of Cobalamin Riboswitches in the Corrinoid-Producing Organohalide Respirer Desulfitobacterium hafniense

“…It would be interesting to test whether Desulfitobacterium-Dehalococcoides cocultures such as those obtained previously (57) would allow the latter to fully dechlorinate tetrachloroethene (PCE) without corrinoid addition in the medium, as was shown in recent studies with Geobacter and other corrinoid-producing bacteria (58,59). Other OHR bacteria such as Sulfurospirillum multivorans (13), Geobacter lovleyi (60), and Dehalobacter restrictus (61) display a large variety of relationships to corrinoid biosynthesis, further emphasizing the importance of studying individual model organisms.…”

“…Unlike in well-known corrinoid-dependent enzymes, the electron-transfer reaction catalyzed by corrinoid-dependent reductive dehalogenases is yet mechanistically unresolved (10)(11)(12). Noteworthy, a new corrinoid, norpseudovitamin B 12 , has been discovered in the tetrachloroethene reductive dehalogenase of Sulfurospirillum multivorans (13). Thus far, only the genomes of D. hafniense strains Y51 (14) and DCB-2 (15) have been published; however, several additional members of this genus are currently part of sequencing projects (http://jgi.doe.gov/).…”

Guided cobalamin biosynthesis supports Dehalococcoides mccartyi reductive dechlorination activity

“…Cobamides with 5 0 ,6 0 -dimethylbenzimidazole (DMB) as the lower a-ligand are cobalamins such as adenosylcobalamin, hydroxocobalamin and cyanocobalamin (CN-Cbl). A cobalamin or a cobamide serve as cofactors for the PCE and TCE reductive dehalogenases (RDases) of several organohalide-respiring genera such as Dehalobacter, Desulfitobacterium and Sulfurospirillum, and exemplify the crucial roles these corrinoids play in the energy metabolism of organohalide-respiring Bacteria [7][8][9][10].…”