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

S. Tsuji, B. M. Martin, D. C. Kaslow, B. R. Migeon, Prabhakara V Choudary, B. K. Stubbleflied, J. A. Mayor, G. J. Murray, J. A. Barranger, E. 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 genoma 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 15 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 - 1987
Externally publishedYes

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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