Native corrinoids from Clostridium cochlearium are adeninylcobamides: spectroscopic analysis and identification of pseudovitamin B(12) and factor A.
The corrinoids from the obligate anaerobe Clostridium cochlearium were extracted as a mixture of Co(beta)-cyano derivatives. From 50 g of frozen cells, approximately 2 mg (1.5 micromol) of B(12) derivatives was obtained as a crystalline sample. Analysis of the corrinoid sample of C. cochlearium by a combination of high-pressure liquid chromatography and UV-Vis absorbance spectroscopy revealed the presence of three cyano corrinoids in a ratio of about 3:1:1. The spectroscopic data acquired for the sample indicated the main components to be pseudovitamin B(12) (Co(beta)-cyano-7"-adeninylcobamide) (60%) and factor A (Co(beta)-cyano-7"-[2-methyl]adeninylcobamide) (20%). Authentic pseudovitamin B(12) was prepared by guided biosynthesis from cobinamide and adenine. Both pseudovitamin B(12) and its homologue, factor A, were subjected to complete spectroscopic analysis by UV-Vis, circular dichroism, mass spectrometry, and by one- and two-dimensional (1)H, (13)C-, and (15)N nuclear magnetic resonance (NMR) spectroscopy. The third component was indicated by the mass spectra to be an isomer of factor A and is likely (according to NMR) to be 7"-[N(6)-methyl]-adeninylcobamide, a previously unknown corrinoid. C. cochlearium thus biosynthesizes as its native "complete" B(12) cofactors the 7"-adeninylcobamides and two homologous corrinoids, in which the nucleotide base is a methylated adenine.
Hoffmann
B
B
Institut für Organische Chemie, Universität Innsbruck, A-6020 Innsbruck, Austria.
Oberhuber
M
M
Stupperich
E
E
Bothe
H
H
Buckel
W
W
Konrat
R
R
Kräutler
B
B
D002942:Circular Dichroism; D003013:Clostridium / Q000737:chemistry* / Q000378:metabolism; D003038:Cobamides / Q000032:analysis*; D045728:Corrinoids; D015394:Molecular Structure; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D011166:Porphyrins / Q000032:analysis*; D013056:Spectrophotometry, Ultraviolet / Q000379:methods; D014805:Vitamin B 12 / Q000031:analogs & derivatives* / Q000032:analysis / Q000096:biosynthesis
D003038:Cobamides; D045728:Corrinoids; D011166:Porphyrins; C413332:factor A; C005602:pseudovitamin B12; C000913:cobamamide; D014805:Vitamin B 12