Electron Paramagnetic Resonance Studies of the Methylmalonyl-CoA Mutase Reaction. Evidence for Radical Intermediates using Natural and Artificial Substrates as well as the Competitive Inhibitor 3-Carboxypropyl-CoA
1994; Wiley; Volume: 225; Issue: 3 Linguagem: Inglês
10.1111/j.1432-1033.1994.0891b.x
ISSN1432-1033
AutoresYmin Zhao, Andreas Abend, Meinrad Kunz, Peter Such, János Rétey,
Tópico(s)Metal-Catalyzed Oxygenation Mechanisms
ResumoThe substrate‐dependent homolysis of the cobalt‐carbon bond and generation of organic radicals in the coenzyme‐B 12 –methylmalonyl‐CoA‐mutase complex have been demonstrated by EPR measurements. Both the natural substrate methylmalonyl‐CoA, its 13 C‐substituted analogue and the non‐hydrolysable synthetic substrates succinyl‐dethia(carba)‐CoA, succinyl‐dethia(dicarba)‐CoA and 4‐carboxy‐2‐oxo‐butyl‐CoA induced similar but not identical EPR signals. 3‐Carboxypropyl‐CoA, a novel competitive inhibitor, has been synthesised. Its K i , value of 89 ± 6 μM was in the same range as the K m , of succinyl‐CoA. Using [5′‐ 3 H]adenosylcobalamin, an enzyme‐dependent tritium transfer to the inhibitor has been shown. The enzyme‐coenzyme‐inhibitor complex also exhibited EPR signals that were less structured and less intensive than the corresponding signals with active substrates. These results prove that the inhibitor also induces cobalt‐carbon bond homolysis and undergoes reversible hydrogen transfer but not rearrangement.
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