Tryptophan fluorescence quenching by alkaline pH and ternary complex formation in human beta 1 beta 1 and horse EE alcohol dehydrogenases. |
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Authors: | T Ehrig B B Muhoberac T D Hurley W F Bosron |
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Institution: | Department of Biochemistry, Indiana University School of Medicine, Indianapolis 46202-5122. |
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Abstract: | The horse EE and human β1β1 alcohol dehydrogenase isoenzymes have almost identical protein backbone folding patterns and contain 2 tryptophans per subunit (Trp-15 and Trp-314). Tyr-286, which had been proposed to quench the fluorescence of Trp-314 by resonance energy transfer at alkaline pH in EE, is substituted by Cys in β1β1. The proposed role of Tyr-286 in pH-dependent quenching of EE is confirmed by our observation that tryptophan fluorescence of β1β1 is not substantially quenched at alkaline pH. Tyr-286 had also been implicated in the quenching of Trp-314 upon formation of the EE-NAD+-trifluoroethanol ternary complex. However, β1β1 exhibits the same extent of tryptophan fluorescence quenching as EE upon complexation, which strongly suggests that Tyr-286 is not involved in ternary complex quenching. |
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Keywords: | Alcohol dehydrogenase Isoenzyme Fluorescence spectroscopy Tryptophan quenching |
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