Spectroscopic Evidence for the Formation of a Four-Coordinate Co²⁺Cobalamin Species upon Binding to the Human ATP:Cobalamin Adenosyltransferase

The human adenosyltransferase hATR converts exogenous cobalamin into coenzyme B₁₂ by transferring the adenosyl group from cosubstrate ATP to a transiently formed Co¹⁺cobalamin (Co¹⁺Cbl) species. A particularly puzzling aspect of hATR function is that the midpoint potential for Co²⁺Cbl → Co¹⁺Cbl reduction is below that of readily available biological reductants. Our magnetic circular dichroism and electron paramagnetic resonance spectroscopic studies reported here reveal that, in the absence of ATP, the interaction between Co²⁺Cbl and hATR promotes partial conversion of the cofactor to its “base-off” form in which a water molecule occupies the lower axial position. This interaction becomes much stronger in the presence of ATP, leading to the formation of an unprecedented Co²⁺Cbl species with spectroscopic signatures consistent with an essentially four-coordinate, square-planar Co²⁺ center. This unusual Co²⁺Cbl coordination is expected to raise the Co^2+/1+ reduction potential well into the physiological range.