Protein-coenzyme interactions in adenosylcobalamin-dependent glutamate mutase.
Glutamate mutase catalyses an unusual isomerization involving free-radical intermediates that are generated by homolysis of the cobalt-carbon bond of the coenzyme adenosylcobalamin (coenzyme B(12)). A variety of techniques have been used to examine the interaction between the protein and adenosylcobalamin, and between the protein and the products of coenzyme homolysis, cob(II)alamin and 5'-deoxyadenosine. These include equilibrium gel filtration, isothermal titration calorimetry, and resonance Raman, UV-visible and EPR spectroscopies. The thermodynamics of adenosylcobalamin binding to the protein have been examined and appear to be entirely entropy-driven, with DeltaS=109 J.mol(-1).K(-1). The cobalt-carbon bond stretching frequency is unchanged upon coenzyme binding to the protein, arguing against a ground-state destabilization of the cobalt-carbon bond of adenosylcobalamin by the protein. However, reconstitution of the enzyme with cob(II)alamin and 5'-deoxyadenosine, the two stable intermediates formed subsequent to homolysis, results in the blue-shifting of two of the bands comprising the UV-visible spectrum of the corrin ring. The most plausible interpretation of this result is that an interaction between the protein, 5'-deoxyadenosine and cob(II)alamin introduces a distortion into the ring corrin that perturbs its electronic properties.
Department of Chemistry, University of Michigan, Ann Arbor, MI 48109-1055, U.S.A.
M S
MS
Huhta
H P
HP
Chen
C
C
Hemann
C R
CR
Hille
E N
EN
Marsh
D003038:Cobamides / Q000378:metabolism*; D004578:Electron Spin Resonance Spectroscopy; D019751:Intramolecular Transferases / Q000378:metabolism*; D013056:Spectrophotometry, Ultraviolet; D013059:Spectrum Analysis, Raman; D013816:Thermodynamics
D003038:Cobamides; D019751:Intramolecular Transferases; C021570:methylaspartate mutase; C000913:cobamamide