Cloning and expression of PTP-PEST. A novel, human, nontransmembrane protein tyrosine phosphatase.
The polymerase chain reaction was used to amplify protein tyrosine phosphatase (PTPase)-related cDNA from a template of total RNA isolated from human skeletal muscle. A novel PTPase, which we term PTP-PEST, was detected by this method. The polymerase chain reaction fragment was used to screen two different HeLa cell libraries to obtain full length cDNA clones. The cDNA predicts a protein of 510 amino acids, approximately 60 kDa, that does not contain an obvious signal sequence or transmembrane segment suggesting it is a nonreceptor type enzyme. The PTPase domain is located in the N-terminal portion of the molecule and displays approximately 35% identity to other members of this family of enzymes. The C-terminal segment is rich in Pro, Glu, Asp, Ser, and Thr residues, possessing features of PEST motifs which have previously been identified in proteins with very short intracellular half-lives. The protein was expressed in Escherichia coli as a fusion product with glutathione S-transferase. Intrinsic activity was demonstrated in vitro against a variety of phosphotyrosine-containing substrates including BIRK, the autophosphorylated cytoplasmic kinase domain of the insulin receptor beta subunit. It did not dephosphorylate phosphoseryl-phosphorylase a. PTP-PEST mRNA is broadly distributed in a variety of cell lines. Stimulation of human rhabdomyosarcoma A204 cells, a transformed muscle line, with insulin led to an approximately 4-fold induction of PTP-PEST mRNA within 36 h.
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Cold Spring Harbor Laboratory, New York 11724.
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Sommercorn
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D000328:Adult; D000595:Amino Acid Sequence; D001483:Base Sequence; D015152:Blotting, Northern; D003001:Cloning, Molecular; D004247:DNA; D006367:HeLa Cells; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D010766:Phosphorylation; D017027:Protein Tyrosine Phosphatases / Q000235:genetics* / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
D004247:DNA; D017027:Protein Tyrosine Phosphatases