Aldehyde dehydrogenase. Maintaining critical active site geometry at motif 8 in the class 3 enzyme. |
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Authors: | J Hempel I Kuo J Perozich B C Wang R Lindahl H Nicholas |
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Affiliation: | Department of Biological Sciences, University of Pittsburgh, Pittsburgh 15620, USA. hempel@psc.edu |
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Abstract: | Alignment of all known, diverse members of the aldehyde dehydrogenase (ALDH) extended family revealed only two strictly conserved, nonglycine residues, a glutamate and a phenylalanine residue. Both occur in one of the highly conserved 'motif' segments and both occupy strategic locations in the tertiary structure at the bottom of the catalytic funnel. In class 3 ALDH, these are Glu333 and Phe335. In addition, Asp247, which is not highly conserved but is characteristic of class 3 ALDHs, hydrogen bonds the main chain between Glu333 and Phe335. These three residues were mutated conservatively. Michaelis constants determined for both NAD/propanal and NADP/benzaldehyde substrate pairs show all three residues to be crucial to effective catalysis, and suggest that the hydrogen bond to Asp247 is a key element in maintaining precise geometry of key elements at the active site. |
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