Identification of a missense mutation in an adult-onset patient with glycogenosis type II expressing only one allele

F. Martiniuk, Mark F. Mehler, M. Bodkin, S. Tzall, K. Hirschhorn, N. Zhong, R. Hirschhorn

Research output: Contribution to journalArticle

21 Citations (Scopus)

Abstract

The lysosomal enzyme acid alpha glucosidase (GAA) or acid maltase is deficient in glycogen storage disease type II. We sought to determine the molecular basis for the disease in an adult-onset patient, unusual for very low enzyme activity similar to that seen with the infantile-onset form and with a previously reported defect in phosphorylation. We constructed cDNA and genomic DNA libraries from the patient's cell line (GM 1935) and determined the nucleotide sequence of the coding region. There were three base-pair substitutions in one allele (C1935 to A; G2446 to A and C2780 to T), all predicting amino acid changes (Asp-645 to Glu; Val-816 to Ile and Thr-927 to Ile). To determine which of the three base-pair substitutions resulted in loss of enzyme activity, we next utilized primer-directed mutagenesis and transient gene expression in an SV40-immortalized GAA- deficient fibroblast cell line. Only the construct containing the G2446 to A mutation (Val-816 to Ile) lost GAA enzyme activity, while the other two substitutions (including the Thr-927 to Ile change that predicts a loss of a potential site for N-linked glycosylation and mannose phosphorylation) each resulted in enzyme activity equal to the control. Analysis of RFLPs in genomic DNA, as well as sequence analysis for the three base-pair alterations, indicated that the patient was a genetic compound. We next digested PCR-amplified cDNA (reverse-transcribed from RNA) with Aat II to detect the base-pair 1935 substitution and found that virtually all of the mRNA was derived from the allele with the three base-pair substitutions. Our results provide the first identification of a disease-specific mutation in one allele of a patient with the adult onset form of glycogenosis type II.

Original languageEnglish (US)
Pages (from-to)681-687
Number of pages7
JournalDNA and Cell Biology
Volume10
Issue number9
StatePublished - 1991

Fingerprint

Glycogen Storage Disease Type II
Missense Mutation
Base Pairing
Alleles
Enzymes
alpha-Glucosidases
Complementary DNA
Phosphorylation
Cell Line
Mutation
Genomic Library
Mannose
Gene Library
Glycosylation
Mutagenesis
Restriction Fragment Length Polymorphisms
Sequence Analysis
Fibroblasts
RNA
Gene Expression

ASJC Scopus subject areas

  • Cell Biology
  • Genetics
  • Molecular Biology

Cite this

Martiniuk, F., Mehler, M. F., Bodkin, M., Tzall, S., Hirschhorn, K., Zhong, N., & Hirschhorn, R. (1991). Identification of a missense mutation in an adult-onset patient with glycogenosis type II expressing only one allele. DNA and Cell Biology, 10(9), 681-687.

Identification of a missense mutation in an adult-onset patient with glycogenosis type II expressing only one allele. / Martiniuk, F.; Mehler, Mark F.; Bodkin, M.; Tzall, S.; Hirschhorn, K.; Zhong, N.; Hirschhorn, R.

In: DNA and Cell Biology, Vol. 10, No. 9, 1991, p. 681-687.

Research output: Contribution to journalArticle

Martiniuk, F, Mehler, MF, Bodkin, M, Tzall, S, Hirschhorn, K, Zhong, N & Hirschhorn, R 1991, 'Identification of a missense mutation in an adult-onset patient with glycogenosis type II expressing only one allele', DNA and Cell Biology, vol. 10, no. 9, pp. 681-687.
Martiniuk, F. ; Mehler, Mark F. ; Bodkin, M. ; Tzall, S. ; Hirschhorn, K. ; Zhong, N. ; Hirschhorn, R. / Identification of a missense mutation in an adult-onset patient with glycogenosis type II expressing only one allele. In: DNA and Cell Biology. 1991 ; Vol. 10, No. 9. pp. 681-687.
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