Isolation and expression in Escherichia coli of a cDNA clone encoding human beta-glucuronidase.
Mucopolysaccharidosis type VII is a lysosomal storage disease resulting from a deficiency of beta-glucuronidase (BG) activity. To facilitate the investigation of mutation in the disease and provide molecular diagnostic tools for affected families, we have isolated human BG cDNA clones. The SV40-transformed human fibroblast cDNA library of Okayama and Berg [Mol. Cell. Biol. 3 (1982) 280-289] was screened with a fragment of a murine BG cDNA clone (pGUS-1). The 17 human cDNA clones (pHUG) isolated were identical by restriction mapping, varying only in length. The pHUG clones show 80% DNA sequence homology with pGUS-1 in a 198-bp PvuII-SstI restriction fragment. Both pGUS-1 and the pHUG clones contained an open reading frame (ORF) throughout the sequenced region with a predicted amino acid sequence homology of 73%. Expression in Escherichia coli of a 1150-bp fragment of pHUG-1 subcloned in pUC9 resulted in an isopropyl-thio-beta-galactoside (IPTG)-inducible 35-kDal fusion protein which was specifically immunoprecipitated by goat anti-human BG immunoglobulin G (IgG). This evidence provides direct confirmation that the pHUG cDNA clones correspond to human BG.
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D004274:DNA, Recombinant; D004926:Escherichia coli / Q000235:genetics; D005786:Gene Expression Regulation; D005966:Glucuronidase / Q000096:biosynthesis / Q000235:genetics*; D006801:Humans; D051379:Mice; D009083:Mucopolysaccharidoses / Q000175:diagnosis / Q000201:enzymology; D010957:Plasmids; D013045:Species Specificity
D004274:DNA, Recombinant; D005966:Glucuronidase