Functional dynamics of response regulators using NMR relaxation techniques.
A fundamental concept of phosphorylation-mediated signaling is the precise switching between discrete functional conformations. According to the traditional view, phosphorylation induces a new, active conformation. In this chapter, a series of NMR experiments performed on a response regulator are described that challenge this traditional notion. The combination of NMR relaxation experiments with chemical shift data and the linkage to structure/function reveals a fundamentally different activation mechanism. The NMR data for the response regulator NtrC provide kinetic (rates of interconversion), thermodynamic (relative populations), and structural (chemical shift) information for the conformational exchange process. The results demonstrate that both the inactive and active states are present before phosphorylation, and activation occurs via a shift of this preexisting equilibrium. This concept is in accordance with the energy landscape view of proteins that embraces the existence of conformational substates. We conjecture that this population-shift mechanism is a general paradigm for response regulator activation and possibly more universal for phosphorylation-mediated signaling.
Gardino
Alexandra K
AK
Department of Biochemistry, Brandeis University, Waltham, MA, USA.
Kern
Dorothee
D
D000495:Allosteric Site; D029968:Escherichia coli Proteins / Q000737:chemistry*; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008956:Models, Chemical; D008958:Models, Molecular; D008968:Molecular Conformation; D051922:PII Nitrogen Regulatory Proteins / Q000737:chemistry*; D010766:Phosphorylation; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D012486:Salmonella typhimurium / Q000378:metabolism; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D013816:Thermodynamics; D013997:Time Factors; D014157:Transcription Factors / Q000737:chemistry*
D029968:Escherichia coli Proteins; D051922:PII Nitrogen Regulatory Proteins; D014157:Transcription Factors; C497876:glnG protein, E coli