Biosynthesis of the corrin macrocycle of coenzyme B12 in Pseudomonas denitrificans.
Studies with cell-free protein preparations from a series of recombinant strains of Pseudomonas denitrificans demonstrated that precorrin-3 is converted into a further trimethylated intermediate, named precorrin-3B, along the pathway to coenzyme B12. It was then shown that the part of the pathway from precorrin-3 (called precorrin-3A hereafter) to precorrin-6x involves three intermediates, precorrin-3B, precorrin-4, and precorrin-5. Precorrin-3B was isolated in its native (reduced) as well as its oxidized (factor-IIIB) states, and precorrin-4 was isolated in its oxidized form only (factor-IV). Both factors were in vitro precursors of precorrin-6x. The synthesis of precorrin-6x from precorrin-3A was shown to be catalyzed by four enzymes, CobG, CobJ, CobM, and CobF, intervening in this order. They were purified to homogeneity. CobG, which converts precorrin-3A to precorrin-3B, was found to be an iron-sulfur protein responsible for the oxidation known to occur between precorrin-3A and precorrin-6x, and CobJ, CobM, and CobF are the C-17, C-11, and C-1 methylases, respectively. The acetate fragment is extruded after precorrin-4 formation. This study combined with our recent structural studies on factor-IV (D. Thibaut, L. Debussche, D. Fréchet, F. Herman, M. Vuilhorgne, and F. Blanche, J. Chem. Soc. Chem. Commun. 1993:513-515, 1993) and precorrin-3B (L. Debussche, D. Thibaut, M. Danzer, F. Debu, D. Fréchet, F. Herman, F. Blanche, and M. Vuilhorgne, J. Chem. Soc. Chem. Commun. 1993:1100-1103, 1993) provides a first step-by-step picture of the sequence of the enzymatic reactions leading to the corrin ring in P. denitrificans.
Debussche
L
L
Département Analyse, Centre de Recherche de Vitry-Alfortville, Rhône-Poulenc Rorer S.A., Vitry-sur-Seine, France.
Thibaut
D
D
Cameron
B
B
Crouzet
J
J
Blanche
F
F
D002247:Carbon Isotopes; D003038:Cobamides / Q000096:biosynthesis*; D045728:Corrinoids; D004926:Escherichia coli / Q000378:metabolism; D007536:Isomerism; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008780:Methyltransferases / Q000378:metabolism; D015394:Molecular Structure; D010957:Plasmids; D011549:Pseudomonas / Q000235:genetics / Q000378:metabolism*; D015183:Restriction Mapping; D012436:S-Adenosylmethionine / Q000378:metabolism; D014805:Vitamin B 12 / Q000737:chemistry / Q000378:metabolism*
D002247:Carbon Isotopes; D003038:Cobamides; D045728:Corrinoids; D012436:S-Adenosylmethionine; D008780:Methyltransferases; C000913:cobamamide; D014805:Vitamin B 12