Identification, characterization, and structure/function analysis of a corrin reductase involved in adenosylcobalamin biosynthesis.
Vitamin B(12), the antipernicious anemia factor, is the cyano derivative of adenosylcobalamin, which is one of nature's most complex coenzymes. Adenosylcobalamin is made along one of two similar yet distinct metabolic pathways, which are referred to as the aerobic and anaerobic routes. The aerobic pathway for cobalamin biosynthesis proceeds via cobalt insertion into a ring-contracted macrocycle, which is closely followed by adenosylation of the cobalt ion. An important prerequisite for adenosylation is the reduction of the centrally chelated metal from Co(II) to a highly nucleophilic Co(I) form. We have cloned a gene, cobR, encoding a biosynthetic enzyme with this co(II)rrin reductase activity from Brucella melitensis. The protein has been overproduced, and the resulting flavoprotein has been purified, characterized, and crystallized and its structure determined to 1.6A resolution. Kinetic and EPR analysis reveals that the enzyme proceeds via a semiquinone form. It is proposed that CobR may interact with the adenosyltransferase to overcome the large thermodynamic barrier required for co(II)rrin reduction.
Lawrence
Andrew D
AD
Protein Science Group, Department of Biosciences, University of Kent, Canterbury, Kent CT2 7NJ, United Kingdom.
Deery
Evelyne
E
McLean
Kirsty J
KJ
Munro
Andrew W
AW
Pickersgill
Richard W
RW
Rigby
Stephen E J
SE
Warren
Martin J
MJ
D017347:Brucella melitensis / Q000201:enzymology; D003001:Cloning, Molecular; D003035:Cobalt / Q000737:chemistry; D003038:Cobamides / Q000096:biosynthesis*; D045728:Corrinoids / Q000737:chemistry*; D018360:Crystallography, X-Ray; D004578:Electron Spin Resonance Spectroscopy; D007477:Ions; D007700:Kinetics; D008956:Models, Chemical; D008968:Molecular Conformation; D010088:Oxidoreductases / Q000737:chemistry* / Q000502:physiology; D013329:Structure-Activity Relationship; D013816:Thermodynamics; D014805:Vitamin B 12 / Q000737:chemistry
D003038:Cobamides; D045728:Corrinoids; D007477:Ions; C473700:corrin; D003035:Cobalt; D010088:Oxidoreductases; C529270:corrin reductase, Brucella melitensis; C000913:cobamamide; D014805:Vitamin B 12