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Krabbe Disease: Isolation and Characterization of a Full-Length cDNA for Human Galactocerebrosidase

1994; Elsevier BV; Volume: 198; Issue: 2 Linguagem: Inglês

10.1006/bbrc.1994.1071

1090-2104

Norio Sakai, Koji Inui, Noriko Fujii, H. Fukushima, Junji Nishimoto, Itaru Yanagihara, Yuji Isegawa, Akihiro Iwamatsu, S Okada,

Trypanosoma species research and implications

Human galactocerebrosidase, the enzyme deficient in Krabbe disease, was purified, through several hydrophobic column steps and gel filtration, 22,650-fold from human lymphocytes. Using information on its N-terminal and internal amino acid sequences, and the polymerase chain reaction method, we cloned a full-length cDNA for the enzyme. The deduced amino acid sequence matched all amino acid sequences determined. The 3780 nucleotide sequence included 2007 nucleotides which encoded a single chain peptide of 669 amino acid residues with a 26 amino acid N-terminal signal peptide and six potential asparagine-linked glycosylation sites. The galactocerebrosidase cDNA detected an about 4 kb mRNA band material in human cultured skin fibroblasts. A nonsense mutation was found at codon 369 (GAA → TAA) in the coding sequence of cDNA amplified from cultured skin fibroblast mRNA from a patient with typical Krabbe disease.