Extended subnanosecond structural dynamics of myoglobin revealed by Laue crystallography

doi:10.1073/pnas.0508880103

Extended subnanosecond structural dynamics of myoglobin revealed by Laue crystallography

*Institut de Biologie Structurale, Unité Mixte de Recherche 5075, Centre National de la Recherche Scientifique/Commissariat à l’Energie Atomique/Université Joseph Fourier, 41 Rue Jules Horowitz, 38027 Grenoble Cedex 1, France;
^†European Synchrotron Radiation Facility, 6 Rue Jules Horowitz, B.P. 220, 38043 Grenoble Cedex, France;
^§Dipartimento di Scienze Biochimiche and Istituto Pasteur-Fondazione Cenci Bolognetti, Università di Roma “La Sapienza,” Piazzale A. Moro 5, 00185 Rome, Italy; and
^¶Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-0520

Edited by Alan R. Fersht, University of Cambridge, Cambridge, United Kingdom, and approved January 31, 2006
↵ ^‡D.B. and B.V. contributed equally to this work. (received for review October 11, 2005)

Abstract

Work carried out over the last 30 years unveiled the role of structural dynamics in controlling protein function. Cavity networks modulate structural dynamics trajectories and are functionally relevant; in globins they have been assigned a role in ligand migration and docking. These findings raised renewed interest for time-resolved structural investigations of myoglobin (Mb), a simple heme protein displaying a photosensitive iron-ligand bond. Photodissociation of MbCO generates a nonequilibrium population of protein structures relaxing over a time range extending from picoseconds to milliseconds. This process triggers ligand migration to matrix cavities with clear-cut effects on the rate and yield of geminate rebinding. Here, we report subnanosecond time-resolved Laue diffraction data on the triple mutant YQR-Mb [Leu-29(B10)Tyr, His-64(E7)Gln, Thr-67(E10)Arg] that depict the sequence of structural events associated with heme and protein relaxation from 100 ps to 316 ns and above. The photodissociated ligand rapidly (<0.1 ns) populates the Xe-binding cavity distal to the heme. Moreover, the heme relaxation toward the deoxy configuration is heterogeneous, with a slower phase (≈ns) evident in these experiments. Damping of the heme response appears to result from a strain exerted by the E-helix via the CD-turn; Phe-43(CD1), in close contact with heme, opposes tilt until the strain is relieved. A comparison with crystallographic data on wild-type Mb and mutants Leu(29)Phe or Leu(29)Trp suggests that the internal structure controls the rate and amplitude of the relaxation events. A correlation between structural dynamics as unveiled by Laue crystallography and functional properties of Mb is presented.

Footnotes

^‖To whom correspondence should be addressed. E-mail: maurizio.brunori{at}uniroma1.it
Author contributions: D.B., B.V., and M.B. designed research; D.B., B.V., A.A., G.S., F.S., and P.A.A. performed research; D.B., B.V., A.A., G.S., F.S., P.A.A., and M.B. analyzed data; and D.B., B.V., P.A.A., and M.B. wrote the paper.
Conflict of interest statement: No conflicts declared.
This paper was submitted directly (Track II) to the PNAS office.
Abbreviations:

Mb,

myoglobin;

wt-Mb,

wild-type Mb;

YQR-Mb,

Mb triple mutant [Leu-29(B10)Tyr, His-64(E7)Gln, Thr-67(E10)Arg];

F-Mb,

Leu-29(B10)Phe Mb mutant;

W-Mb,

Leu-29(B10)Trp Mb mutant;

MD,

molecular dynamics.