Functional and molecular modelling studies of two hereditary fructose intolerance-causing mutations at arginine 303 in human liver aldolase

摘要

pWe have identified a novel hereditary fructose intolerance mutation in the aldolase B gene (i.e. liver aldolase) that causes an arginine-to-glutamine substitution at residue 303 (Argsup303/sup → Gln). We previously described another mutation (Argsup303/sup → Trp) at the same residue. We have expressed the wild-type protein and the two mutated proteins and characterized their kinetic properties. The catalytic efficiency of protein Glnsup303/sup is approx. 1/100 that of the wild-type for substrates fructose 1,6-bisphosphate and fructose 1-phosphate. The Trpsup303/sup enzyme has a catalytic efficiency approx. 1/4800 that of the wild-type for fructose 1,6-bisphosphate; no activity was detected with fructose 1-phosphate. The mutation Argsup303/sup → Trp thus substitution impairs enzyme activity more than Argsup303/sup → Gln. Three-dimensional models of wild-type, Trpsup303/sup and Glnsup303/sup aldolase B generated by homology-modelling techniques suggest that, because of its larger size, tryptophan exerts a greater deranging effect than glutamine on the enzyme9s three-dimensional structure. Our results show that the Argsup303/sup → Gln substitution is a novel mutation causing hereditary fructose intolerance and provide a functional demonstration that Argsup303/sup, a conserved residue in all vertebrate aldolases, has a dominant role in substrate binding during enzyme catalysis./p