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Minimize RSR Award Detail

Research Spending & Results

Award Detail

Awardee:RUTGERS, THE STATE UNIVERSITY OF NEW JERSEY
Doing Business As Name:Rutgers University New Brunswick
PD/PI:
  • Charalampos Kalodimos
  • (848) 445-6560
  • babis@rutgers.edu
Award Date:07/25/2011
Estimated Total Award Amount: $ 622,186
Funds Obligated to Date: $ 622,186
  • FY 2011=$207,143
  • FY 2012=$207,822
  • FY 2013=$207,221
Start Date:08/01/2011
End Date:07/31/2014
Transaction Type:Grant
Agency:NSF
Awarding Agency Code:4900
Funding Agency Code:4900
CFDA Number:47.074
Primary Program Source:040100 NSF RESEARCH & RELATED ACTIVIT
Award Title or Description:Structural and Dynamic Studies of Catabolite Activator Protein Complexes
Federal Award ID Number:1121896
DUNS ID:001912864
Parent DUNS ID:001912864
Program:Molecular Biophysics

Awardee Location

Street:33 Knightsbridge Road
City:Piscataway
State:NJ
ZIP:08854-3925
County:Piscataway
Country:US
Awardee Cong. District:06

Primary Place of Performance

Organization Name:Rutgers University New Brunswick
Street:33 Knightsbridge Road
City:Piscataway
State:NJ
ZIP:08854-3925
County:Piscataway
Country:US
Cong. District:06

Abstract at Time of Award

Intellectual merit Genetic regulatory proteins target specific sites within the genome and either enhance or repress transcriptional activity to elicit cellular responses. The Escherichia coli catabolite activator protein (CAP; referred to also as the cAMP receptor protein, CRP) is a universal transcriptional activator that regulates the expression of over two hundred genes. CAP has long served as the textbook example for understanding transcription regulation. CAP has provided a classic model system for structural and mechanistic studies of transcription activation. Mechanistic descriptions of transcription activation, developed for CAP, are more nearly complete than descriptions of any other examples of transcription activation. Nevertheless, the complete structural basis for CAP-mediated transcription activation remains unknown. Notably, over the recent years CAP has provided an excellent system in which to examine the structure- and dynamics-function relationships that form the basis of allostery. CAP has provided the first experimentally identified system wherein allosteric interactions are mediated through changes in protein motions, in the absence of changes in the mean structure of the protein. The main objectives of this project are to use CAP as a model system to address fundamental questions regarding allosteric regulation and transcriptional activation. An integrated structural, dynamic, and thermodynamic approach will be used to (1) characterize the dynamics of CAP mutants with altered allosteric properties and their interaction energetics with DNA; (2) determine the solution structure of the class I and class II CAP-dependent promoter subassemblies and (3) determine the structural basis for the assembly of the entire CAP-mediated transcription initiation complex. Broader impact In addition to addressing fundamental biological questions, this project will be used to train students in structural biology, biophysics, and molecular biology, areas that are rapidly becoming integrated in 21st century science. Postdocs, graduate and undergraduate students will have the opportunity to be involved in a multi-disciplinary project that aims at the development of groundbreaking methodologies to enable the structural and dynamic characterization of supramolecular protein complexes by high resolution NMR spectroscopy. This will enable researchers to approach problems from a multidisciplinary and interactive perspective, thus experiencing first hand the utility of applying state-of-the-art methodologies to important biological problems. The paradigm of combining structural, dynamic, thermodynamic and kinetic approaches to study complex protein systems will be included in a new course, currently designed by the PI, to exemplify the value of using an interdisciplinary and quantitative approach to answer questions of scientific importance. The course is intended for a large, diverse audience consisting of graduate and advanced undergraduate students in the programs of Molecular Biosciences, Chemistry and Chemical Biology, Biomedical Engineering and BIOMAPS at Rutgers University.

Publications Produced as a Result of this Research

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Tzeng, Shiou-Ru and Kalodimos, Charalampos G "{Protein activity regulation by conformational entropy.}" Nature, v.488, 2012, p.236--240.

Tinberg, Christine E and Khare, Sagar D and Dou, Jiayi and Doyle, Lindsey and Nelson, Jorgen W and Schena, Alberto and Jankowski, Wojciech and Kalodimos, Charalampos G and Johnsson, Kai and Stoddard, Barry L and Baker, David "{Computational design of ligand-binding proteins with high affinity and selectivity.}" Nature, v.501, 2013, p.212--216.

Kalodimos, Charalampos G "{NMR reveals novel mechanisms of protein activity regulation.}" Protein science : a publication of the Protein Society, v.20, 2011, p.773--782.

S-R. Tzeng and C.G. Kalodimos "Protein activity regulation by conformational entropy" Nature, v.488, 2012, p.236-240. doi:doi:10.1038/nature11271 

Tzeng, Shiou-Ru and Kalodimos, Charalampos G "{Allosteric inhibition through suppression of transient conformational states.}" Nature Chemical Biology, v.9, 2013, p.462--465.

Kalodimos, Charalampos G "{Protein function and allostery: a dynamic relationship.}" Ann N Y Acad Sci, v.1260, 2012, p.81--86.

Tzeng, Shiou-Ru and Kalodimos, Charalampos G "{Protein dynamics and allostery: an NMR view.}" Current opinion in structural biology, v.21, 2011, p.62--67.


Project Outcomes Report

Disclaimer

This Project Outcomes Report for the General Public is displayed verbatim as submitted by the Principal Investigator (PI) for this award. Any opinions, findings, and conclusions or recommendations expressed in this Report are those of the PI and do not necessarily reflect the views of the National Science Foundation; NSF has not approved or endorsed its content.

Accumulating evidence indicates that internal dynamics can mediate allosteric interactions and modulate ligand binding. It is now well accepted that it is the inextricable link between structure and dynamics that ultimately control protein activity. However, how the interplay between protein structure and internal dynamics regulates protein function is poorly understood. Often, ligand binding, post-translational modifications and mutations modify protein activity in a manner that is not possible to rationalize solely on the basis of structural data. Although internal motions of proteins appear to have a major role in regulating protein activity, the nature of their contributions remains elusive, especially in quantitative terms. Over the last few years, our lab has reported novel and exciting findings about protein activity regulatory mechanisms through modulation of internal dynamics on different timescales. We have exploited the favorable properties of the catabolite activator protein (CAP), a prototype for allosterically regulated proteins, and used atomic- resolution NMR spectroscopy to characterize the structure and dynamics of several CAP variants with modified allosteric features. We have shown that changes in conformational entropy can determine whether protein–ligand interactions will occur, even among protein complexes with identical binding interfaces. We have further shown that that allosteric inhibition can be effected by destabilizing a low-populated conformational state that serves as an on-pathway intermediate for ligand binding, without altering the protein’s ground-state structure. The findings are of extreme importance as they, collectively, demonstrate how changes in fast internal dynamics (conformational entropy) and slow internal dynamics (energetically excited conformational states) can regulate binding activity in a way that cannot be predicted on the basis of the protein’s ground-state structure. 

In this project we used an integrated structural, dynamic and thermodynamic approach to study biological questions of fundamental interest and importance. Postdocs, graduate and undergraduate students that participate in the project were involved in the development of the methodologies that enabled characterization of large protein complexes by high resolution NMR spectroscopy. They were also involved in studying dynamic properties and understanding the often elusive role of protein dynamics in protein function. The complimentary techniques enabled our people to approach problems from a multidisciplinary and interactive perspective, thus experiencing first hand the utility of applying state-of- the-art methodologies to important biological problems.

 


Last Modified: 08/01/2014
Modified by: Charalampos Kalodimos

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