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Crystal structure of an aldehyde reductase Y50F mutant‐NADP complex and its implications for substrate binding
Author(s) -
Ye Qilu,
Hyndman David,
Green Nancy,
Li Xuhong,
Korithoski Bryan,
Jia Zongchao,
Flynn T. Geoffrey
Publication year - 2001
Publication title -
proteins: structure, function, and bioinformatics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.699
H-Index - 191
eISSN - 1097-0134
pISSN - 0887-3585
DOI - 10.1002/prot.1066
Subject(s) - cofactor , ternary complex , stereochemistry , aldehyde , chemistry , aldehyde reductase , tyrosine , active site , binding site , enzyme , mutant , substrate (aquarium) , reductase , oxidoreductase , phenylalanine , biochemistry , catalysis , biology , amino acid , ecology , gene
Pig aldehyde reductase containing the active site mutation tyrosine(50) to phenylalanine has been crystallized in the presence of the cofactor NADP(H) to a resolution of 2.2 Å. This structure clearly shows loss of the tyrosine hydroxyl group and no other significant perturbations compared with previously determined structures. The mutant binds cofactor (both oxidized and reduced) more tightly than the wild‐type enzyme but shows a complete lack of binding of the aldehyde reductase inhibitor barbitone, as determined by fluorescence titrations. Numerous attempts at preparing a ternary complex with a range of small aldehyde substrates were unsuccessful. This result, in addition to the inability of the mutant protein to bind the inhibitor, provides strong evidence for the proposal that the tyrosine hydroxyl group is essential for substrate binding in addition to catalysis. Proteins 2001;44:12–19. © 2001 Wiley‐Liss, Inc.