Sequential dephosphorylation of a multiply phosphorylated insulin receptor peptide by protein tyrosine phosphatases

Ramachandran, C., Aebersold, R., Tonks, N. K., Pot, D. A. (May 1992) Sequential dephosphorylation of a multiply phosphorylated insulin receptor peptide by protein tyrosine phosphatases. Biochemistry, 31 (17). pp. 4232-4238. ISSN 0006-2960

URL: http://www.ncbi.nlm.nih.gov/pubmed/1373652
DOI: 10.1021/bi00132a012

Abstract

The question of whether protein tyrosine phosphatases (PTPases) dephosphorylate a multiply phosphorylated peptide in a random or ordered manner was investigated using the synthetic triphosphotyrosyl peptide TRDIY(P)ETDY(P)Y(P)RK, corresponding to the major sites of autophosphorylation of the insulin receptor, as a substrate for four purified PTPases. All four enzymes dephosphorylated the triphospho peptide to produce diphospho, monophospho, and nonphosphorylated forms. Partially dephosphorylated peptides were separated by reverse-phase HPLC, and the di- and monophospho peptides were collected and analyzed by solid-phase sequencing to determine the order of dephosphorylation of the three sites by each of the PTPases. The quantitative analysis of the signals for derivatives of tyrosine and phosphotyrosine generated at Positions 5, 9, and 10 of the peptide showed that the low molecular weight human placental PTPase 1B preferentially dephosphorylated the two phosphotyrosines at positions 9 and 10 whereas the integral membrane enzyme CD45 (from human spleen) and the bacterially expressed rat LAR preferentially dephosphorylated the phosphotyrosine at position 5. A second low molecular weight enzyme, termed TCPTPase, did not display any specificity for a particular phosphotyrosyl residue. These results demonstrate that different PTPases exhibit a characteristic pattern of dephosphorylation of the triphospho peptide model substrate, raising the possibility that features in the primary structure surrounding the dephosphorylation site may contribute to substrate specificity.

Item Type: Paper
Uncontrolled Keywords: HUMAN-PLACENTA BETA-SUBUNIT SIGNAL TRANSDUCTION CLONING FAMILY KINASE AUTOPHOSPHORYLATION EXPRESSION PURIFICATION MEMBER
Subjects: bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein types > enzymes
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein types > insulin receptor
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein expression > phosphorylation
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein expression
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein types
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein types > enzymes > protein tyrosine phosphatase
CSHL Authors:
Communities: CSHL labs > Tonks lab
Depositing User: Matt Covey
Date: May 1992
Date Deposited: 17 Dec 2013 19:32
Last Modified: 17 Dec 2013 19:32
Related URLs:
URI: http://repository.cshl.edu/id/eprint/29081

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