Multiple forms of the human tyrosine phosphatase RPTP alpha. Isozymes and differences in glycosylation.
Among all the receptor-linked protein-tyrosine-phosphatase RPTP alpha clones described from mammalian tissues, one differed in that it encoded a 9-amino-acid insert 3 residues upstream from the transmembrane segment (Kaplan, R., Morse, B., Huebner, K., Croce, C., Howk, R. Ravera, M., Ricca, G., Jaye, M., and Schlessinger, J. (1990) Proc. Natl. Acad. Sci. U. S. A. 87, 7000-7004). Using the polymerase chain reaction technique, simultaneous expression of both isoforms was demonstrated in human T-cell and vascular smooth muscle libraries, as well as in the A431 human epidermal cancer cell line. Following transient expression in COS-1 cells, each isoform gave rise to two proteins of 100 and 130 kDa, respectively. Endoglycosidase treatment showed that the 100-kDa species corresponded to a molecule exclusively glycosylated on N-residues, whereas the 130-kDa species contained both, N- and O-linked carbohydrates. Pulse-chase experiments demonstrated that the smaller RPTP alpha protein is a precursor of the larger one. A high affinity antibody was generated that recognizes the immature protein only; however, both proteins can be detected by Western blot analysis after a simple chemical hydrolysis. Following Superose 12 chromatography, the 100- and 130-kDa species of RPTP alpha emerged as 200- and 340-kDa proteins, respectively. Both species exhibited similar enzymatic activities as determined with a peptide substrate in immunoprecipitates.
Daum
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Department of Biochemistry SJ-70, University of Washington, Seattle 98195.
Regenass
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Sap
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Schlessinger
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J
Fischer
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EH
D000595:Amino Acid Sequence; D001483:Base Sequence; D002460:Cell Line; D018076:DNA, Complementary; D004792:Enzyme Precursors / Q000235:genetics / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017027:Protein Tyrosine Phosphatases / Q000235:genetics / Q000378:metabolism*; D054630:Receptor-Like Protein Tyrosine Phosphatases, Class 4; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D018076:DNA, Complementary; D004792:Enzyme Precursors; D007527:Isoenzymes; D011956:Receptors, Cell Surface; C516841:PTPRA protein, human; D017027:Protein Tyrosine Phosphatases; D054630:Receptor-Like Protein Tyrosine Phosphatases, Class 4