Different Reaction Mechanisms for Mesophilic and Thermophilic Dihydrofolate Reductases

摘要

Dihydrofolate reductase (DHFR) is a ubiquitous enzyme that catalyzes the reduction of 7,8-dihydrofolate (H_2F) to 5,6,7,8-tetrahydrofolate (H_4F) using NADPH as a cofactor. During the DHFR-catalyzed reaction the pro-R hydrogen on C4 of NADPH is transferred to C6 of H_2F with concomitant protonation of N5 of H_2F. Hydride transfer catalyzed by mesophilic DHFR from Escherichia coli (EcDHFR) and hyperthermophilic DHFR from Thermotoga mariiima (TmDHFR) shows a sigmoidal pH profile with pK_a 6.5 and 6.0, respectively. The pK_a of the EcDHFR-catalyzed reaction is due to protonation of the substrate itself rather than the active-site Asp27 residue.