S. Koster et al., LOCATION OF THE ACTIVE-SITE AND PROPOSED CATALYTIC MECHANISM OF PTERIN-4A-CARBINOLAMINE DEHYDRATASE, European journal of biochemistry, 241(3), 1996, pp. 858-864
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 t
he mutants showed reduced activity, and only the Glu57-->Ala mutant an
d the His61-->Ala. His62-->Ala double mutant were fully devoid of acti
vity. The dissociation constants of quinonoid 6,6-dimethyl-7,8-dihydro
pterin were significantly increased for binding to the Glu57-->Ala, Hi
s61-->Ala. His62-->Ala single mutants and the His61-->Ala, His62-->Ala
double mutant, confirming that His61 and His62 are essential for subs
trate binding and catalysis. The mechanism of dehydration is proposed
to involve base catalysis at the N(5)-H group of the substrate by His6
1.