LSIII, a long neurotoxin

Proteins are not static; they undergo internal motions spanning many orders of magnitude in rate and amplitude. The importance of fluctuations for biological function is becoming increasingly clear. NMR is a powerful probe of dynamics, reporting on both the extent and timescale of internal motions. We are characterizing the structure and dynamics of LSIII, a neurotoxin that binds selectively to the nicotinic acetylcholine receptor (AChR). We discovered that the principal binding loop, which appears disordered in crystal structures of related proteins, exhibits considerable local order, apparently undergoing rigid-body motion about a hinge region relative to the protein core. This has implications for the thermodynamics of AChR recognition by LSIII, and provides clues to the architecture of the binding site on AChR. The loop dynamics appear to enable LSIII to reach around an occlusion at the entrance to the binding site, which could be a mechanism used by the receptor to discriminate between large and small molecules, or to ensure that the toxin remains bound to the receptor as it undergoes binding-induced conformational change. In addition to studies of magnetic relaxation (using 13C), we are performing theoretical investigations of LSIII dynamics (using MD simulation).
    Molecular Dynamics of LSIII, Peter J. Connolly, Alan S. Stern, Christopher J. Turner, and Jeffrey C. Hoch, Biochemistry 42, 14443-51 (2003)
    The Solution Structure of LSIII, a Long Neurotoxin from the Venom of Laticauda semifasciata, Peter J. Connolly, Alan S. Stern, and Jeffrey C. Hoch, Biochemistry 35, 418-426 (1996)
    Solution Structure of LSSIII, With Possible ossible Implications for Binding to the Acetylcholine Receptor, Peter J. Connolly, Alan S. Stern, and Jeffrey C. Hoch, in "Dynamics and the Problem of Recognition in Biological Macromolecules", O. Jardetzky and J.-F. Lefevre, eds., Plenum Press, New York (1996)

Aims: We have overexpressed LSIII in E. coli, which will enable us to introduce stable isotopes and to study dynamical and thermodynamic properties of mutant proteins. We will perform additional MD simulations to characterize the free energy profile of the binding loop dynamics.