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Science 291 (5512): 2429-2433

Copyright © 2001 by the American Association for the Advancement of Science

Two-State Allosteric Behavior in a Single-Domain Signaling Protein

Brian F. Volkman,1* Doron Lipson,2 David E. Wemmer,3 Dorothee Kern2dagger

Protein actions are usually discussed in terms of static structures, but function requires motion. We find a strong correlation between phosphorylation-driven activation of the signaling protein NtrC and microsecond time-scale backbone dynamics. Using nuclear magnetic resonance relaxation, we characterized the motions of NtrC in three functional states: unphosphorylated (inactive), phosphorylated (active), and a partially active mutant. These dynamics are indicative of exchange between inactive and active conformations. Both states are populated in unphosphorylated NtrC, and phosphorylation shifts the equilibrium toward the active species. These results support a dynamic population shift between two preexisting conformations as the underlying mechanism of activation.

1 National Magnetic Resonance Facility at Madison (NMRFAM), Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA.
2 Department of Biochemistry, Brandeis University, Waltham, MA 02454, USA.
3 Physical Biosciences Division, Lawrence Berkeley National Laboratory and Department of Chemistry, University of California, Berkeley, CA 94720, USA.
*   Present address: Department of Biochemistry, Medical College of Wisconsin, Milwaukee, WI 53226, USA.

dagger    To whom correspondence should be addressed. E-mail: dkern{at}brandeis.edu

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