Molecular cloning and expression of cDNAs encoding human alpha-mannosidase II and a previously unrecognized alpha-mannosidase IIx isozyme.
Golgi alpha-mannosidase II (alpha-MII) is an enzyme involved in the processing of N-linked glycans. Using a previously isolated murine cDNA clone as a probe, we have isolated cDNA clones encompassing the human alpha-MII cDNA open reading frame and initiated isolation of human genomic clones. During the isolation of genomic clones, genes related to that encoding alpha-MII were isolated. One such gene was found to encode an isozyme, designated alpha-MIIx. A 5-kb cDNA clone encoding alpha-MIIx was then isolated from a human melanoma cDNA library. However, comparison between alpha-MIIx and alpha-MII cDNAs suggested that the cloned cDNA encodes a truncated polypeptide with 796 amino acid residues, while alpha-MII consists of 1144 amino acid residues. To reevaluate the sequence of alpha-MIIx cDNA, polymerase chain reaction (PCR) was performed with lymphocyte mRNAs. Comparison of the sequence of PCR products with the alpha-MIIx genomic sequence revealed that alternative splicing of the alpha-MIIx transcript can result in an additional transcript encoding a 1139-amino acid polypeptide. Northern analysis showed transcription of alpha-MIIx in various tissues, suggesting that the alpha-MIIx gene is a housekeeping gene. COS cells transfected with alpha-MIIx cDNA containing the full-length open reading frame showed an increase of alpha-mannosidase activity. The alpha-MIIx gene was mapped to human chromosome 15q25, whereas the alpha-MII gene was mapped to 5q21-22.
Glycobiology Program, La Jolla Cancer Research Foundation, CA 92037, USA.
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Misago
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Liao
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Kudo
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Eto
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Mattei
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Moremen
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Fukuda
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D001483:Base Sequence; D002874:Chromosome Mapping; D002884:Chromosomes, Human, Pair 15; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D015723:Gene Library; D015894:Genome, Human; D006801:Humans; D007527:Isoenzymes / Q000096:biosynthesis / Q000235:genetics*; D008214:Lymphocytes / Q000201:enzymology; D008361:Mannosidases / Q000096:biosynthesis / Q000235:genetics*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000096:biosynthesis; D012641:Selection, Genetic; D017386:Sequence Homology, Amino Acid; D014018:Tissue Distribution
D018076:DNA, Complementary; D007527:Isoenzymes; D011994:Recombinant Proteins; D008361:Mannosidases; C038829:mannosyl-oligosaccharide 1,3 - 1,6-alpha-mannosidase