Vibrational echo studies of protein dynamics

Dana D. Dlott (Curriculum Vitae)

Dana D. Dlott (personal home page)

David M. Dlott (personal home page)

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This project involves time-resolved coherent infrared (IR) spectroscopy of ligands such as CO bound to the active sites of proteins. Macromolecular vibrational spectra are usually broad and poorly resolved, due to vibrational congestion (see Myoglobin spectrum at right). Nuclear magnetic resonance (NMR) spectra of macromolecules are similarly congested, but in recent years powerful coherent multidimensional pulse sequences have been developed to overcome that difficulty. With recent advances in coherent IR pulse generation, similar pulsed and coherent IR techniques can now be applied to biomolecules.

The simplest coherent pulse sequence is the vibrational echo experiment, analogous to the spin echo. The vibrational echo removes inhomogeneous broadening to reveal the underlying homogeneous vibrational transition. By combining echoes with IR pump-probe techniques, which measures the vibrational lifetime, vibrational pure dephasing processes can be studied in proteins for the first time. Pure dephasing is sensitive to fluctuations in protein structure occurring on the time scale of 0.1 to 100 ps.

These experiments are performed at the Stanford Free-electron Laser Center. So far we have used this new laser source to study native heme proteins, mutant proteins made by site-directed mutagenesis, and totally synthetic heme model compounds.

Most of our studies have concentrated on the myoglobin (Mb) protein. In Mb, the active site is located on a prosthetic group, the aromatic molecule protoheme. Our most significant finding is that the heme group detects fluctuations of protein structure and transmits them to the ligand at the active site. By looking at these fluctuation rates as a function of temperature, we have seen a very surprising feature of proteins -- the fluctuation rates vary with temperature in the same way as various glass systems. The rate is proportional to temperature raised to the 1.3 power. However this T1.3 behavior in glasses vanishes when the temperature is raised to only 10K, whereas in Mb, it can be observed extending all the way to room temperature. This surprising finding provides deep insights into the hierarchical dynamics of complex macromolecular systems.

To view a presentation on vibrational echoes in heme proteins, click here.

REPRESENTATIVE PUBLICATIONS

"Mutant and wild type myoglobin-CO protein dynamics: vibrational echo experiments", K. D. Rector, C. W. Rella, J. R. Hill, A. S. Kwok, S. G. Sligar, E. Y. T. Chien, Dana D. Dlott and M. D. Fayer, J. Phys. Chem. 101, pp. 1468-1475 (1997).

"Dynamics of myoglobin-CO with the proximal histidine removed: vibrational echo experiments", Kirk D. Rector, J. R. Engholm, J. R. Hill, D. J. Myers, R. Hu, S. G. Boxer, Dana D. Dlott and M. D. Fayer, J. Phys. Chem. B. 102 pp. 331 (1998).

wpe1.jpg (9964 bytes)
Structure of the heme protein Myoglobin (Mb)

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Infrared spectrum of Mb-CO. The sharp peak at right is due to the CO stretching transition

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Free-electron laser at Stanford University

Mbechodecay.gif (16980 bytes)

ViVibrational echo decay of CO bound to Mb at 80K

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TTemperature dependent pure dephasing for Mb-CO and a mutant with distal histidine removed