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Epimerase-deficiency galactosemia is the result of impairment of the human enzyme UDP-galactose-4-epimerase (hGALE). Substitution mutations have been identified in the hGALE alleles of patients with the clinically mild, peripheral epimerase deficiency. The first identification of an hGALE mutation in a patient with the clinically severe, generalized epimerase deficiency has been made. V94M was found on both GALE alleles.

One of the fundamental questions concerning expression and function of dimeric enzymes involves the impact of naturally occurring mutations on subunit assembly and heterodimer activity. This question is of particular interest for the human enzyme galactose-1-phosphate uridylyl-transferase (GALT), impairment of which results in the inherited metabolic disorder galactosemia, because many if not most patients studied to date are compound heterozygotes rather than true molecular homozygotes. Furthermore, the broad range of phenotypic severity observed in these patients raises the possibility that allelic combination, not just allelic constitution, may play some role in determining outcome. In the work described herein, we have selected two distinct naturally occurring null mutations of GALT, Q188R and R333W, and asked the questions (i) what are the impacts of these mutations on subunit assembly, and (ii) if heterodimers do form, are they active? To answer these questions, we have established a yeast system for the coexpression of epitope-tagged alleles of human GALT and investigated both the extent of specific GALT subunit interactions and the activity of defined heterodimer pools. We have found that both homodimers and heterodimers do form involving each of the mutant subunits tested and that both heterodimer pools retain substantial enzymatic activity. These results are significant not only in terms of their implications for furthering our understanding of galactosemia and GALT holoenzyme structure - function relationships but also because the system described may serve as a model for similar studies of other complexes composed of multiple subunits.

The cloning ofa human galactokinase gene from the complementation of a galactokinase-deficient strain of Saccharomyces cerevisiae was reported. The cDNA is predicted to encode a protein of 458 amino acids. The galactokinase gene was mapped to chromosome 15. However, a previous studies gave mapped another human galactokinase gene to chromosome 17. It is therefore unknown which of the two galactokinase gene loci are affected in galactokinase deficiency disease.

A study was conducted on the molecular characterization of L&4P and F171S galactosemia mutations and S135L polymorphism present in human galactose-1-phosphate uridyltransferase (GALT). Results show that the galactosemia mutations decrease enzymatic activity while the polymorphism has near normal activity. Both mutations indicated functional importance based on their influence in evolutionary conserved residues. On the other hand, the polymorphism was found to occur in a nonconserved domain. The mutations and polymorphism proved effective in the study of human GALT functional domains.