Galactosemia: A strategy to identify new biochemical phenotypes and molecular genotypes

Journal Article · Wed Mar 01 04:00:00 EST 1995 · American Journal of Human Genetics

OSTI ID:70440

Elsas, L J; Langley, S; Steele, E; Evinger, J; Brown, A; Singh, R; Fernhoff, P; Hjelm, L N; Dembure, P P; Fridovich-Keil, J L ^[1]

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1. Emory Univ. School of Medicine, Atlanta, GA (United States)

We describe a stratagem for identifying new mutations in the galactose-1-phosphate uridyl transferase (GALT) gene. GALT enzyme activity and isoforms were defined in erythrocytes from probands and their first-degree relatives. If the biochemical phenotypes segregated in an autosomal recesssive pattern, we screened for common mutations by using multiplex PCR and restriction endonuclease digestions. If common mutant alleles were not present, the 11 exons of the GALT gene were amplified by PCR, and variations from the normal nucleotide sequences were identified by SSCP. The suspected region(s) was then analyzed by direct DNA sequencing. We identified 86 mutant GALT alleles that reduced erythrocyte GALT activity. Seventy-five of these GALT genomes had abnormal SSCP patterns, of which 41 were sequenced, yielding 12 new and 21 previously reported, rare mutations. Among the novel group of 12 new mutations, an unusual biochemical phenotype was found in a family whose newborn proband has classical galactosemia. He had inherited two mutations in cis (N314D-E204K) from his father, whose GALT activity was near normal, and an additional GALT mutation in the splice-acceptor site of intron C (IVSC) from his mother. The substitution of a positively charged E204K mutation created a unique isoform-banding pattern. An asymptomatic sister`s GALT genes carries three mutations (E203K-N314D/N314D) with eight distinct isoform bands. Surprisingly, her erythrocytes have normal GALT activity. We conclude that the synergism of pedigree, biochemical, SSCP, and direct GALT gene analyses is an efficient protocol for identifying new mutations and speculate that E203K and N314D codon changes produce intra-allelic complementation when in cis. 40 refs., 4 figs., 3 tabs.

• OSTI ID: 70440
• Journal Information: American Journal of Human Genetics, Journal Name: American Journal of Human Genetics Journal Issue: 3 Vol. 56; ISSN AJHGAG; ISSN 0002-9297
• Country of Publication: United States
• Language: English

Related Subjects

55 BIOLOGY AND MEDICINE
BASIC STUDIES
BIOCHEMISTRY
DIAGNOSIS
DNA SEQUENCING
ENZYMES
GALACTOSE
GENE MUTATIONS
GENES
GENOME MUTATIONS
GENOTYPE
HEREDITARY DISEASES
METABOLISM
PATIENTS
PHENOTYPE
POLYMERASE CHAIN REACTION