Gated and ungated electron transfer reactions from aromatic amine dehydrogenase to azurin

Interprotein electron transfer (ET) occurs between the tryptophan tryptophy lquinone (TTQ) prosthetic group of aromatic amine dehydrogenase (AADH) and copper of azurin, The ET reactions from two chemically distinct reduced for ms of TTQ were studied: an O-quinol form that was generated by reduction by dithionite, and an N-quinol form that was generated by reduction by substr ate. It was previously shown that on reduction by substrate, an amino group displaces a carbonyl oxygen on TTQ, and that this significantly alters the rate of its oxidation by azurin (Hyun, Y-L., and Davidson V, L, (1995) Bio chemistry 34, 12249-12254). To determine the basis for this change in react ivity, comparative kinetic and thermodynamic analyses of the ET reactions f rom the O-quinol and N-quinol forms of TTQ in AADH to the copper of azurin were performed. The reaction of the O-quinol exhibited values of electronic coupling (H-AB) Of 0.13 cm(-1) and reorganizational energy (lambda) of 1.6 eV, and predicted an ET distance of approximately 15 Angstrom, These resul ts are consistent with the ET event being the rate-determining step for the redox reaction, Analysis of the reaction of the N-quinol by Marcus theory yielded an H-AB,which exceeded the nonadiabatic limit and predicted a negat ive ET distance. These results are diagnostic of a gated ET reaction. Solve nt deuterium kinetic isotope effects of 1.5 and 3.2 were obtained, respecti vely, for the ET reactions from O-quinol and N-quinoI AADH indicating that transfer of an exchangeable proton was involved in the rate-limiting reacti on step which gates ET from the N-quinol, but not the O-quinol, These resul ts are compared with those for the ET reactions from another TTQ enzyme, me thylamine dehydrogenase, to amicyanin, The mechanism by which the ET reacti on of the N-quinol is gated is also related to mechanisms of other gated in terprotein ET reactions.