Bovine Serum Albumin-Catalyzed Deprotonation of [1-13C]Glycolaldehyde: Protein Reactivity toward Deprotonation of the α-Hydroxy α-Carbonyl Carbon

Abstract

Bovine serum albumin (BSA) in D₂O at 25 °C and pD 7.0 was found to catalyze the deuterium exchange reactions of [1-¹³C]glycolaldehyde ([1-¹³C]GA) to form [1-¹³C,2-²H]GA and [1-¹³C,2,2-di-²H]GA. The formation of [1-¹³C,2-²H]GA and [1-¹³C,2,2-di-²H]GA in a total yield of 51 ± 3% was observed at early reaction times, and at later times, [1-¹³C,2-²H]GA was found to undergo BSA-catalyzed conversion to [1-¹³C,2,2-di-²H]GA. The overall second-order rate constant for these deuterium exchange reactions [(k_E)_P] equals 0.25 M⁻¹ s⁻¹. By comparison, (k_E)_P values of 0.04 M⁻¹ s⁻¹ [Go, M. K., Amyes, T. L., and Richard, J. P. (2009) Biochemistry 48, 5769−5778] and 0.06 M⁻¹ s⁻¹ [Go, M. K., Koudelka, A., Amyes, T. L., and Richard, J. P. (2010) Biochemistry 49, 5377−5389] have been determined for the wild-type- and K12G mutant TIM-catalyzed deuterium exchange reactions of [1-¹³C]GA, respectively, to form [1-¹³C,2,2-di-²H]GA. These data show that TIM and BSA exhibit a modest catalytic activity toward deprotonation of the α-hydroxy α-carbonyl carbon. We suggest that this activity is intrinsic to many globular proteins, and that it must be enhanced to demonstrate meaningful de novo design of protein catalysts of proton transfer at α-carbonyl carbon.