Vibrational Probe and Methods Development for Studying the Ultrafast Dynamics of Preferential Solvation of Biomolecules by 2D-IR.

Abstract

Over the last decade two-dimensional infrared spectroscopy (2D-IR) has emerged as a powerful method for the investigation of biological samples and their dynamics. Through the implementation of state of the art signal processing methods we have demonstrated a significant, 20-fold, reduction in the acquisition time of traditional 2D-IR spectra. This new technique, utilizing compressed sensing, compliments our previously developed RASD method, allowing for the rapid acquisition of complete 2D-IR spectra as opposed to dynamical information at a single excitation-detection frequency pair. Additionally we have realized the first biocompatible, modular, metal-carbonyl probes for 2D-IR utilizing benzyl-chromium tribarbonyls. This has enabled ultrafast 2D-IR investigations of lipids and preferential solvation in solutions and at site-specific locations within enzyme scaffolds. In aqueous solutions we find that preferential solvation by a polar cosolvent causes a slowdown of the observed dynamics sensed by our probes. From modeling our system this slowdown is found to be consistent with arising from the slow, ca. 8 ps, exchange dynamics between the polar co-solute and water in the vicinity of our probe. This interpretation of preferential solvation in solution is further able to describe the observed dynamical differences found at the protein-solvent interface in a model system. By studying a series of protein mutants we find, spectroscopically and through simulations, that interactions between the side chains and the solution are sufficient to modulate the degree of preferential solvation and therefore dynamics, within specific sites of the protein. This information provides a foundation on how to modulate of the diffusion of substrates and products into and out-of the active sites of enzymes, through directed mutation of their protein sequence. The diffusional motion of the solvent and substrates is often the rate-limiting step in enzymatic catalysis. By controlling the local solvation dynamics of enzymes, sequence mutations offer a method to fine-tune the dynamics of enzymes. The ability to characterize the site-specific solvation dynamics of enzymes in response to primary structure mutations, positions 2D-IR and our chromium tricarbonyl probes as powerful tools for understanding protein and enzyme dynamics. This provides insight into controlling the catalytic rate of enzymes through directed mutation.