Quenching of the intrinsic fluorescence of liver alcohol dehydrogenase by the alkaline transition and by coenzyme binding.

摘要: The fluorescence of alcohol dehydrogenase is quenched by the acid dissociation some group on protein having an apparent pKa 9.6 at 25 degrees C. this alkaline quenching transition unchanged binding trifluoroethanol or pyrazole to enzyme selective removal active site Zn2+ ion. This indicates that ionization a zinc-bound water molecule not responsible for quenching. NAD+ causes drop in and shift lower pH. In ternary complex formed with difficult discern between pH 6 11. NAD+-pyrazole complex, however, small but noticeable observed pKa(app) approximately 9.5. We propose centered shifted upon NAD+. Instead, amplitude effect decreased point it detect when bound. present model describes dependence concentration terms two independently operating, dynamic mechanisms. Our data cast serious doubt identification, made previously literature, whose coupled binding.(ABSTRACT TRUNCATED AT 250 WORDS)