Premium
Thermosensitive mutants of the MPTP and hPTP1B protein tyrosine phosphatases: Isolation and structural analysis
Author(s) -
Muise Eric S.,
Vrielink Alice,
Ennis Maurice A.,
Lemieux Nancy H.,
Tremblay Michel L.
Publication year - 1996
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1002/pro.5560050405
Subject(s) - thermostability , mutant , mutagenesis , biochemistry , cysteine , enzyme , site directed mutagenesis , tyrosine , chemistry , phosphatase , protein tyrosine phosphatase , mutation , microbiology and biotechnology , biology , gene
A PCR‐based random mutagenesis procedure was employed to identify several thermosensitive mutants of the MPTP enzyme, the murine homologue of the human T‐cell PTPase and rat PTP‐S enzymes. Four mutants with varying degrees of thermosensitivity were characterized for their thermostability and refolding properties following incubation at the nonpermissive temperature. Structure analysis of these mutations based on the hPTP1B coordinate structure demonstrates a clear relationship between the position of each mutated amino acid relative to the catalytic cysteine residue and their thermostability. Introduction of two of these mutations in the related enzyme hPTP1B suggests that the structural defects and the resulting thermosensitivity of these mutations may represent an intrinsic property of all PTPase catalytic domains.