• Research Scientist
  • Senior Principal Engineer
  • Western Digital Corporation
  • 44100 Osgood Rd.
  • Fremont, CA 94539
  • lfchen02@yahoo.com

I was a post doc in Professor Taylor’s lab from 1997-1999. I came from a material science background. This was my first appointment into a structural biology laboratory. My work involved building and reforming atom models into 3-D volumes obtained by electron tomography of insect flight muscle in rigor.

Lattice constraints as well as differences in biochemical state of the independently acting myosin heads result in structural variations in situ that depart significantly from the uniform structure that is obtained by X-ray crystal structures. Our aim is to fit initially the atomic model for rigor acto-S1 complex into the envelope of 3-D reconstructions obtained by electron tomography. Adjustments are then made to the atomic model manually using the crystallography program “O” to correct for regions where model and envelope differ. Adjustments in the atomic model at this point are restricted to the light chain domain, which is pivoted about residue 770 of the myosin heavy chain. The motor domain and the actin monomers are kept constant. We have developed a procedure for correcting poor contacts that are produced by these manual adjustments using the RSref program. RSref is a real space refinement program that compares the electron density of the model with the density map. Movements of the model are determined using conjugate gradients. An independent scale factor is used to account for overall differences in density between model and map. The TNT refinement program is used in parallel to determine the corrections needed to improve poor geometry. A set of initial constraints is used to limit movements of the model. One of these involves maintaining the C-terminal residues of the two heavy chains to a minimum separation. Results obtained by modeling the acto-S1 structure into the envelope of the 2-headed crossbridge in rigor insect flight muscle show a range of axial movement in the light chain domain comparable to those hypothesized for muscle contraction. In addition, azimuthal movements are required to bring the two heavy chains into proximity.

Figure Legend. This picture shows an atomic model of an opposed pair of 2-headed myosin crossbridges attached to an actin filament. The model has been fit to an electron density map of insect flight muscle in the rigor state. The 3-D map was obtained by electron tomography of a section of plastic embedded tissue. The model was developed starting with the coordinates of rigor acto-S1 developed by Ivan Rayment and coworkers. We then moved the C-terminal residues to within 12Å of each other by pivoting the light chain domains of both S1s about residue 770. Then RSref was run to correct poor geometry and iverlaping domains. The rsulting model fits the electron density much better than the original starting model which had considerable density falling outside of the 3-D envelope. This project done in collaboration with the laboratories of Michael Chapman at Florida State University and Michael and Mary Reedy at Duke University.

 

PUBLICATIONS

  • L.F. Chen, M. S. Chapman, E. Blanc and K. A. Taylor. Modeling the atomic structure of myosin S1 and actin into 3-D reconstructions of insect flight muscle. Biophys. J. 74, A22 (1998)
  • Kenneth A. Taylor, Holger Schmitz, Mary C. Reedy, Yale E. Goldman, Clara Franzini-Armstrong, Hiro Sasaki, Richard T. Tregear, Kate Poole, Carmen Lucaveche, Robert J. Edwards, Li Fan Chen, Hanspeter Winkler, and Michael K. Reedy. Tomographic 3-D reconstruction of quick frozen, Ca++-activated contracting insect flight muscle. Cell 99, 421-431 (1999).
  • Li Fan Chen, Eric Blanc, Michael S. Chapman and Kenneth A. Taylor. Real space refinement of acto-myosin structures from sectioned muscle. J. Struct. Biol. 133(2), 221-232 (2001)
  • Chen, Li Fan, Winkler, Hanspeter, Reedy, Michael K., Reedy, Mary C. & Taylor ,Kenneth A.. Molecular Modeling of Averaged Rigor Crossbridges from Tomograms of Insect Flight Muscle. J. Struct. Biol., 138(2), 92-104 (2002)
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