Primary structure and processing of the Candida tsukubaensis alpha-glucosidase. Homology with the rabbit intestinal sucrase-isomaltase complex and human lysosomal alpha-glucosidase.
The nucleotide sequence of a 4.39-kb DNA fragment encoding the alpha-glucosidase gene of Candida tsukubaensis is reported. The cloned gene contains a major open reading frame (ORF 1) which encodes the alpha-glucosidase as a single precursor polypeptide of 1070 amino acids with a predicted molecular mass of 119 kDa. N-terminal amino acid sequence analysis of the individual subunits of the purified enzyme, expressed in the recombinant host Saccharomyces cerevisiae, confirmed that the alpha-glucosidase precursor is proteolytically processed by removal of an N-terminal signal peptide to yield the two peptide subunits 1 and 2, of molecular masses 63-65 kDa and 50-52 kDa, respectively. Both subunits are secreted by the heterologous host S. cerevisiae in a glycosylated form. Coincident with its efficient expression in the heterologous host, the C. tsukubaensis alpha-glucosidase gene contains many of the canonical features of highly expressed S. cerevisiae genes. There is considerable sequence similarity between C. tsukubaensis alpha-glucosidase, the rabbit sucrase-isomaltase complex (proSI) and human lysosomal acid alpha-glucosidase. The cloned DNA fragment from C. tsukubaensis contains a second open reading frame (ORF 2) which has the capacity to encode a polypeptide of 170 amino acids. The function and identity of the polypeptide encoded by ORF 2 is not known.
Kinsella
B T
BT
Guinness Brewing Worldwide Research Centre, St. James's Gate Brewery, Dublin, Ireland.
Hogan
S
S
Larkin
A
A
Cantwell
B A
BA
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002175:Candida / Q000201:enzymology*; D003001:Cloning, Molecular; D004271:DNA, Fungal; D004792:Enzyme Precursors / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005087:Glucan 1,4-alpha-Glucosidase / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D007422:Intestines / Q000201:enzymology*; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010957:Plasmids; D011499:Protein Processing, Post-Translational; D011817:Rabbits; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics; D012689:Sequence Homology, Nucleic Acid; D013394:Sucrase-Isomaltase Complex / Q000235:genetics* / Q000378:metabolism; D000520:alpha-Glucosidases / Q000235:genetics* / Q000378:metabolism
D004271:DNA, Fungal; D004792:Enzyme Precursors; D013394:Sucrase-Isomaltase Complex; D000520:alpha-Glucosidases; D005087:Glucan 1,4-alpha-Glucosidase