Immediate objective of this proposal is to describe the particle of tobacco mosaic virus (TMV) and other helical viruses, including the distantly related TMV strain cucumber green mottle mosaic virus (watermelon strain) and the unrelated tobacco rattle virus (TRV), in molecular detail. This forms part of a broader objective: to study the process of macromolecular assembly; in particular, the assembly of macromolecular structures involving proteins and nucleic acids. The present model of TMV has been refined against data at 2.9A resolution to an R factor of 0.10, using the retrained least squares procedure of Hendrickson and Konnert, adapted for fiber diffraction data. Difference Fourier maps have been used to correct the model and locate water molecules. This refinement will continue, and we will continue to improve the difference Fourier technique, which we have recently adapted to take into consideration the special problems of fiber diffraction. The refined model will be used to further elucidate the mechanism of assembly of TMV. Specimens of TMV at low pH will be set up and diffraction patterns recorded. Electron density maps of TMV at low pH will directly reveal the interactions between carboxyl groups from different subunits. These interactions provide a switch, which is used in the assembly and disassembly of the virus. X-ray fiber diffraction studies of CGMMV-W will continue. We plan to search for heavy-atom derivatives in addition to the recently found lead derivative, and to use these derivatives to begin the process of phase determination and calculation of an electron density map, initially at low resolution. Stocks will be built up of the TMV strain U2, for similar work in the future. We plan to improve the quality of our TRV diffraction patterns, by varying the solution conditions under which specimens for X- ray diffraction experiments are made.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM033265-09
Application #
3282728
Study Section
Biophysical Chemistry Study Section (BBCB)
Project Start
1983-09-01
Project End
1993-03-31
Budget Start
1991-04-01
Budget End
1993-03-31
Support Year
9
Fiscal Year
1991
Total Cost
Indirect Cost
Name
Vanderbilt University Medical Center
Department
Type
Schools of Arts and Sciences
DUNS #
004413456
City
Nashville
State
TN
Country
United States
Zip Code
37212
Wang, H; Planchart, A; Allen, D et al. (1993) Preliminary X-ray diffraction studies of ribgrass mosaic virus. J Mol Biol 234:902-4
Pattanayek, R; Stubbs, G (1992) Structure of the U2 strain of tobacco mosaic virus refined at 3.5 A resolution using X-ray fiber diffraction. J Mol Biol 228:516-28
Pattanayek, R; Elrod, M; Stubbs, G (1992) Characterization of a putative calcium-binding site in tobacco mosaic virus. Proteins 12:128-32
Lyne, J E; Carter, D C; He, X M et al. (1990) Preliminary crystallographic examination of a novel fungal lysozyme from Chalaropsis. J Biol Chem 265:6928-30
Lobert, S; Stubbs, G (1990) Fiber diffraction analysis of cucumber green mottle mosaic virus using limited numbers of heavy-atom derivatives. Acta Crystallogr A 46 ( Pt 12):993-7
Stubbs, G (1989) The probability distributions of X-ray intensities in fiber diffraction: largest likely values for fiber diffraction R factors. Acta Crystallogr A 45 ( Pt 3):254-8
Namba, K; Pattanayek, R; Stubbs, G (1989) Visualization of protein-nucleic acid interactions in a virus. Refined structure of intact tobacco mosaic virus at 2.9 A resolution by X-ray fiber diffraction. J Mol Biol 208:307-25
Jeng, T W; Crowther, R A; Stubbs, G et al. (1989) Visualization of alpha-helices in tobacco mosaic virus by cryo-electron microscopy. J Mol Biol 205:251-7
Namba, K; Caspar, D L; Stubbs, G (1988) Enhancement and simplification of macromolecular images. Biophys J 53:469-75
Lobert, S; Heil, P D; Namba, K et al. (1987) Preliminary X-ray fiber diffraction studies of cucumber green mottle mosaic virus, watermelon strain. J Mol Biol 196:935-8

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