Long-Range Protein Vibrations Dependence on Ligand Binding: Rate Promoting Motions

Abstract

It has been suggested that long-range intramolecular vibrations in enzymes may provide efficient access to intermediate state configurations, enhancing catalytic turnover rates. Recently we have successfully measured these long-range protein vibrations using an anisotropic THz near-field technique to measure protein crystals, called crystal anisotropy terahertz microscopy (CATM). The method isolates these motions from the isotropic librational background, revealing narrow band resonances. Recent measurements on free chicken-egg white lysozyme (CEWL) and tri-N-acetylglucosamine (3NAG) inhibitor bound CEWL reveal dramatic and reproducible changes in the intramolecular vibrational mode spectra with binding. A large resonance at ∼70 cm−1 for free CEWL, is somewhat suppressed with binding, whereas a new resonance is observed at ∼ 40 cm−1, with binding. This dramatic change in the spectra with binding confirms that the observed resonances arise from intramolecular vibrations, and not crystal phonons. Using normal mode analysis, our calculated CATM spectra find a consistent increase in the low frequency signal with binding. While the density of states does not change appreciably with binding, the overall direction of motion shifts for vibrations in the frequency range where there is a large change in the optical signal. The change in the direction of the vibrational motion results in a change in the motion of the charge distribution and therefore, the dipole derivative, leading to the sensitivity to the trajectories in the terahertz optical absorbance. The remaining question is if these specific motions participate in the structural preorganization of the binding site, and thus promote catalytic activity. We will discuss measurements to answer this long standing question. This work was supported by NSF MRI∧2 grant DBI295998.