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Free keywords:
hepatocyte nuclear factor-1α-dimerization cofactor; dehydratase; tetrahydrobiopterin; phenyl-alanine hydroxylase
Abstract:
Based on the recently solved three-dimensional structure of pterin-4a-carbinolamine dehydratase from rat/human liver the involvement of the proposed active-site residues Glu57, Asp60, His61, His62, Tyr69, His79, Arg87 and Asp88 was examined by site-directed mutagenesis. Most of the mutants showed reduced activity, and only the Glu57→Ala mutant and the His61→Ala, His62→Ala double mutant were fully devoid of activity. The dissociation constants of quinonoid 6,6-dimethyl-7,8-dihydropterin were significantly increased for binding to the Glu57→Ala, His61→Ala, His62→Ala single mutants and the His61→Ala, His62→Ala double mutant, confirming that His61 and His62 are essential for substrate binding and catalysis. The mechanism of dehydration is proposed to involve base catalysis at the N(5)-H group of the substrate by His61.
