Signal sequence and DNA‐mediated expression of human lysosomal α‐galactosidase A

Shoji TSUJI, Brian M. MARTIN, David C. KASLOW, Barbara R. MIGEON, Prabhakara V. CHOUDARY, Barbara K. STUBBLEFLIED, June A. MAYOR, Gary J. MURRAY, John A. BARRANGER, Edward I. GINNS

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Abstract

Twelve complementary DNA clones for human lysosomal α‐galactosidase A were isolated from an Okayama‐Berg library constructed from SV40‐transformed human fibroblasts. The identity of these clones was confirmed by complete colinearity of the nucleotide‐deduced amino acid sequence with that determined by direct chemical sequencing of human placental α‐galactosidase A. Hybridization of the α‐galactosidase A cDNA to genomic DNA from individuals with varying numbers of X chromosomes as well as from interspecies somatic‐cell hybrids showed only a single locus in the genome at Xq 13.1 – Xq 22. One cDNA clone (pcD‐AG210) contained the complete coding sequence for both the signal peptide and mature α‐galactosidase A. The signal peptide of 31 amino acids contains the expected hydrophobic domains consisting of Leu‐Gly‐Cys‐Ala‐Leu‐Ala‐Leu and Phe‐Leu‐Ala‐Leu‐Val and has Ala at the signal peptidase cleavage site. Twelve out of fifteen G residues flanking the 5′ end of the cDNA in pcD‐AG210 were removed and the truncated fragment was ligated into the original vector. This construct, pcD‐AG502, encoded enzymatically active human α‐galactosidase A in monkey COS cells.

Original languageEnglish (US)
Pages (from-to)275-280
Number of pages6
JournalEuropean Journal of Biochemistry
Volume165
Issue number2
DOIs
StatePublished - Jun 1987

ASJC Scopus subject areas

  • Biochemistry

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    TSUJI, S., MARTIN, B. M., KASLOW, D. C., MIGEON, B. R., CHOUDARY, P. V., STUBBLEFLIED, B. K., MAYOR, J. A., MURRAY, G. J., BARRANGER, J. A., & GINNS, E. I. (1987). Signal sequence and DNA‐mediated expression of human lysosomal α‐galactosidase A. European Journal of Biochemistry, 165(2), 275-280. https://doi.org/10.1111/j.1432-1033.1987.tb11438.x