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Reversible dissociation and unfolding of the dimeric protein thymidylate synthase
Author(s) -
Perry Kathy M.,
Pookanjanatavip Manee,
Zhao Jia,
Santi Daniel V.,
Stroud Robert M.
Publication year - 1992
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.5560010611
Subject(s) - thymidylate synthase , chemistry , dissociation (chemistry) , atp synthase , biophysics , biochemistry , stereochemistry , enzyme , crystallography , biology , genetics , fluorouracil , chemotherapy
Abstract Conditions for in vitro unfolding and refolding of dimeric thymidylate synthase from Lactobacillus casei were found. Ultraviolet difference and circular dichroism spectra showed that the enzyme was completely unfolded at concentrations of urea over 5.5 M. As measured by restoration of enzyme activity, refolding was accomplished when 0.5 M potassium chloride was included in the refolding mixture. Recombination of subunits from catalytically inactive mutant homodimers to form an active hybrid dimer was achieved under these unfolding–refolding conditions, demonstrating a monomer to dimer association step.