Functional characterization of the K257R and G319E-hGALE alleles found in patients with ostensibly peripheral epimerase deficiency galactosemia.

Epimerase deficiency galactosemia is an autosomal recessive condition resulting from the impairment of UDP-galactose 4'-epimerase (hGALE). Although a small number of clinically severe patients have been reported who exhibit "generalized" GALE deficiency, the vast majority exhibit an apparently benign "peripheral" form of the disorder in which enzyme impairment is restricted to the circulating red and white blood cells. Previously, preliminary data were reported suggesting that GALE deficiency is 10-fold more common among African-Americans than among non-African-Americans, and that two missense mutations, K257R and G319E, are found in at least some of these patients. We report here functional studies of these alleles involving expression of the substituted human enzymes in a null-background strain of yeast. Although under normal assay conditions both substituted proteins demonstrate enzyme activities indistinguishable from the wild-type, one (G319E) demonstrates mild impairment under conditions of substrate limitation. No impairments are evident under conditions of cofactor (NAD) limitation. These results are consistent with the apparently benign status of peripheral epimerase deficiency galactosemia, but leave open the question of why patients with these substitutions demonstrate GALE deficiency in their red blood cells. While the possibility remains that K257R and G319E may cause tissue-specific impairments not recapitulated in vitro or in yeast, an equally if not more plausible explanation suggested by interspecies sequence alignments is that both substitutions may be polymorphisms that exist in linkage disequilibrium with other, as yet unidentified causal mutations.