您的账号已在其他设备登录，您当前账号已强迫下线， 如非您本人操作，建议您在会员中心进行密码修改 确定Biochimica et biophysica acta>Kinetic studies on the binding of 1,N6-etheno-NAD+ to glutamate dehydrogenase from Clostridium symbiosum. Kinetic studies on the binding of 1,N6-etheno-NAD+ to glutamate dehydrogenase from Clostridium symbiosum. The mechanism of the binding of reduced coenzyme (NAD+) to clostridial glutamate dehydrogenase (GDH) was determined by transient kinetics. The fluorescent 1,N6-ethenoadenine analogue of NAD+ (epsilonNAD+) was used as a probe of nucleotide binary and ternary complex formation because the binding of NAD+ is optically silent. The kinetics of epsilonNAD+ binding were consistent with a 3-step binding process. The enzyme was found to oscillate between two conformational forms, termed E1 and E2, in the presence and absence of L-glutamate. However, L-glutamate shifted the equilibrium from 96.8% to 99% of the enzyme in the E1 form. The rapid-equilibrium binding of epsilonNAD+ to the E2 form was rate limited by a slow isomerisation of the ternary complex as the binary complex became saturated with epsilonNAD+. The L-glutamate binary complex had a greater affinity for the coenzyme (Kd = 11 microM) than the free enzyme (Km = 39 microM), indicative of a positive interaction of the substrate and coenzyme binding sites. Steady-state studies were also indicative of a positive interaction in the formation of the catalytic complex, with this complex having a Kd for epsilonNAD+ of 6.8 microM. Consequently, there is stabilization of successive complexes on the reaction pathway. 更多 Krebs Institute for Biomolecular Research, Department of Molecular Biology and Biotechnology, University of Sheffield, Western Bank, UK. 特别提示：本网站仅提供医学学术资源服务，不销售任何药品和器械，有关药品和器械的销售信息，请查阅其他网站。