title: Anaerobic chlorophyll isocyclic ring formation in Rhodobacter capsulatus requires a cobalamin cofactor.
abstract: The isocyclic ring of bacteriochlorophyll (BChl) is formed by the conversion of Mg-protoporphyrin monomethyl ester (MPE) to protochlorophyllide (PChlide). Similarities revealed by blast searches with the putative anaerobic MPE-cyclase BchE suggested to us that this protein also uses a cobalamin cofactor. We found that vitamin B(12) (B(12))-requiring mutants of the bluE and bluB genes of Rhodobacter capsulatus, grown without B(12), accumulated Mg-porphyrins. Laser desorption/ionization time-of-flight (LDI-TOF) MS and NMR spectroscopy identified them as MPE and its 3-vinyl-8-ethyl (mvMPE) derivative. An in vivo assay was devised for the cyclase converting MPE to PChlide. Cyclase activity in the B(12)-dependent mutants required B(12) but not protein synthesis. The following reaction mechanism is proposed for this MPE-cyclase reaction. Adenosylcobalamin forms the adenosyl radical, which leads to withdrawal of a hydrogen atom and formation of the benzylic-type 13(1)-radical of MPE. Withdrawal of an electron gives the 13(1)-cation of MPE. Hydroxyl ion attack on the cation gives 13(1)-hydroxy-MPE. Withdrawal of three hydrogen atoms leads successively to 13(1)-keto-MPE, its 13(2)-radical, and cyclization to PChlide.
authors.affiliation: Departments of Physiology and Chemistry, Carlsberg Laboratory, Gamle Carlsberg Vej 10, 2500 Valby, Denmark. gough@biobase.dk
authors.forename: S P
authors.initials: SP
authors.lastname: Gough
authors.forename: B O
authors.initials: BO
authors.lastname: Petersen
authors.forename: J O
authors.initials: JO
authors.lastname: Duus
mesh_terms: D000595:Amino Acid Sequence; D000693:Anaerobiosis; D001429:Bacteriochlorophylls / Q000378:metabolism*; D002855:Chromatography, Thin Layer; D003038:Cobamides / Q000502:physiology; D007501:Iron / Q000502:physiology; D008969:Molecular Sequence Data; D009154:Mutation; D010105:Oxygenases; D011166:Porphyrins / Q000378:metabolism; D016354:Rhodobacter capsulatus / Q000235:genetics / Q000378:metabolism*; D014805:Vitamin B 12 / Q000502:physiology*
chemical_list: D001429:Bacteriochlorophylls; D003038:Cobamides; D011166:Porphyrins; D007501:Iron; D010105:Oxygenases; C053161:magnesium protoporphyrin monomethyl ester oxidative cyclase; C000913:cobamamide; D014805:Vitamin B 12