J Biol Chem, Vol. 273, Issue 15, 8958-8964, April 10, 1998
Kinetic and Thermodynamic Studies of Purine Repressor Binding
to Corepressor and Operator DNA
Han
Xu,
Markos
Moraitis,
Ross J.
Reedstrom, and
Kathleen S.
Matthews
From the Department of Biochemistry & Cell Biology, Rice
University, Houston, Texas 77005
The kinetic and thermodynamic parameters for
purine repressor (PurR)-operator and PurR-guanine binding were
determined using fluorescence spectroscopy and nitrocellulose filter
binding. Operator binding affinity was increased by the presence of
guanine as demonstrated previously (Choi, K. Y., Lu, F., and
Zalkin, H. (1994) J. Biol. Chem. 269, 24066-24072;
Rolfes, R. J., and Zalkin, H. (1990) J. Bacteriol.
172, 5637-5642), and conversely guanine binding affinity was
increased by the presence of operator. Guanine enhanced operator affinity by increasing the association rate constant and decreasing the
dissociation rate constant for binding. Operator had minimal effect on
the association rate constant for guanine binding; however, this DNA
decreased the dissociation rate constant for corepressor by ~10-fold.
Despite significant sequence and structural similarity between PurR and
LacI proteins, PurR binds to its corepressor ligand with a lower
association rate constant than LacI binds to its inducer ligand.
However, the rate constant for PurR-guanine binding to operator is
~3-fold higher than for LacI binding to its cognate operator under
the same solution conditions. The distinct metabolic roles of the
enzymes under regulation by these two repressor proteins provide a
rationale for the observed functional differences.
Copyright © 1998 by The American Society for Biochemistry and Molecular Biology, Inc.
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