The Energetics and Mechanistic Roles of Protein Conformational Dynamics
The Energetics and Mechanistic Roles of Protein Conformational Dynamics
Haw Yang
Department of Chemistry, the University of California at Berkeley, Berkeley, California 94720, USA
Many enzymes mould their structures to poise substrates in their active sites for chemical reaction such that conformational remodeling is necessary during each catalytic cycle. In both protein tyrosine phosphatise B, PtpB, from M. tuberculosis (a virulence factor of M. tuberculosis) and adenylate kinase, AK, from E. coli (a ubiquitous energy-balancing enzyme in cells), the conformational change involves large-amplitude rearrangements of the enzyme’s lid domain. These domain movements have been followed in real time on their respective catalytic timescales using high-resolution single-molecule Förster resonance energy transfer (FRET) spectroscopy. It is shown quantitatively that both PtpB and AK are capable of dynamically sampling two distinct states that correlate well with those observed by x-ray crystallography. Surprisingly, the equilibrium favors the closed, active-site-forming configurations even in the absence of substrates, contrasting the widely accepted induced-fit picture. For PtpB, the results suggest a “dynamic filtering” mechanism by which PtpB has evolved to utilize thermal fluctuations for substrate recognition and for active site protection in the highly oxidative macrophage interior. For AK, the experiments further showed that interaction with substrates restricts the spatial extent of conformational fluctuations rather than locking the enzyme into a compact state. Integrating these microscopic dynamics into macroscopic kinetics allows us to model AK’s lid opening-coupled product release as the enzyme's rate-limiting step—direct evidence for conformation-gated enzymatic reactivity. These experiments suggest that dynamics is an integral component in evolutionary conservation (AK) and adaptation (PtpB)—a concept that may open up new ways of engineering enzymes by controlling the dynamics.
