Progress in FY2013 was in the following areas: (1) HIV-1 CAPSID ASSEMBLIES. We have acquired a large set of 2D and 3D solid state NMR spectra of tubular CA assemblies, with both uniformly 15N,13C-labeled CA and with partially labeled CA, grown on selectively labeled glycerol media. From these spectra, and using computational methods developed in our group, we have determined site-specific assignments of the majority of 15N and 13C NMR chemical shifts for tubular CA assemblies. These assignments allow us to distinguish and identify rigid and mobile segments of the CA sequence within tubular assemblies. Quantitative measurements of 15N-15N dipole-dipole couplings, performed with """"""""backbone recoupling"""""""" methods developed in our group, allow us to identify conformational changes that accompany assembly of CA from its soluble, dimeric form to the hexagonal lattice of the tubes. Solid state NMR spectra also shed light on intermolecular interactions and the molecular structural basis for curvature of the hexagonal lattice. These results will be submitted for publication before the end of FY13. (2) MEMBRANE-ASSOCIATED HIV-1 MATRIX PROTEIN. We have performed initial solid state NMR measurements to assess the feasibility of studies to investigate intermolecular interactions of membrane-bound HIV-1 MA proteins. Solid state NMR spectra show that HIV-1 MA is rather loosely bound to phospholipid bilayer surfaces under experimental conditions used in previous solution NMR studies. We are now investigating the dependence of MA binding on lipid composition and other factors, both to provide new information about interactions that affect lipid binding and to optimize conditions for solid state NMR measurements. (3) HEPATITIS B VIRUS CAPSID STRUCTURE. In collaboration with Dr. Norman Watts, we have acquired 2D and 3D solid state NMR spectra of uniformly 15N,13C-labeled HBV capsid protein, assembled into capsid particles with icosahedral symmetry. Analysis of the data is in progress. We expect to obtain new information about site-specific dynamics and site-specific structural variations that underlie quasi-equivalence in T=3 and T=4 capsid particles.

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