Cloning and characterization of the human gene encoding aspartyl beta-hydroxylase.
Sequence information for aspartyl beta-hydroxylase (AspH), which specifically hydroxylates one Asp or Asn residue in certain epidermal growth factor (EGF)-like domains of a number of proteins, is so far only described for bovine species. We have isolated a 4.3-kb cDNA encoding the human AspH (hAspH) by immunoscreening of a human osteosarcoma (MG63) cDNA library in lambda ZAP with an antiserum raised against membrane fractions of these cells. Northern blot analyses revealed two transcripts with lengths of 2.6 and 4.3 kb. The deduced amino acid (aa) sequence of this cDNA encodes a protein of 757 aa (85 kDa). Comparison with the deduced bovine AspH (bAspH) aa sequence showed striking differences in the N-terminal portion of this protein. In vitro transcription and translation in the presence of canine pancreas microsomes yielded a 56-kDa protein. Western blot analyses of membrane fractions from MG63 cells with AspH-specific antibodies revealed a protein of the same M(r). These results suggest a posttranslational cleavage of the catalytic C terminus in the lumen of the endoplasmic reticulum.
Korioth
F
F
Universität Bielefeld, Fakultät für Chemie, Germany.
Gieffers
C
C
Frey
J
J
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D001859:Bone Neoplasms; D002417:Cattle; D002462:Cell Membrane / Q000201:enzymology; D003001:Cloning, Molecular; D004815:Epidermal Growth Factor / Q000378:metabolism; D015186:Hominidae / Q000235:genetics*; D006801:Humans; D007106:Immune Sera; D006899:Mixed Function Oxygenases / Q000096:biosynthesis / Q000235:genetics*; D008969:Molecular Sequence Data; D012516:Osteosarcoma; D014176:Protein Biosynthesis; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
D007106:Immune Sera; D004815:Epidermal Growth Factor; D006899:Mixed Function Oxygenases; C058291:aspartic acid 2-oxoglutarate-dependent dioxygenase