This project uses image processing techniques to analyze electron micrographs. To answer important questions in structural biology, it is necessary to obtain relatively high resolution 2- and 3-D structural information about biological macromolecules. A specific focus of our research is the understanding of viral structures. At present we are continuing our efforts to investigate the structure of a large animal virus, human herpes simplex virus (type 1). Using the 3-D icosahedral reconstruction technique, we make use of the symmetry of these virus particles to both find the orientation of randomly oriented capsid particles (in ice) and to combine many particles into a 3-D reconstruction. Biological material for these herpes virus reconstructions is provided through a collaboration with researchers at the University of Virginia, Charlottesville and from the Upjohn Co., Kalamazoo. The electron microscopy is performed in LSB, NIAMS. Interpretation of our 3-D reconstruction is performed jointly by all collaborators. 3-D reconstructions of bacculovirus-produced herpes proteins (from Upjohn) have been used to produce reconstituted capsids (at the University of Virginia), which we have shown are identical to herpes virus produced in vivo. By systematically combining different proteins, we have obtained additional information about the high resolution details of herpes virus capsid structure which shows that VP26, a small 12 Kda protein is found on the hexons, but not on the pentons of the capsid. This result has been submitted for publication. In another related research project, we have analyzed the structure of the channel catfish virus, a HSV-1 cousin which turns out to be amazingly similar to it. We may be able to learn more about the evolution of herpes virus capsids from such studies. Future work on this project involves the use of additional antibodies to confirm our localization experiments of other major proteins, and a continuing attempt to increase the resolution of our results substantially. The computational demands of the 3-D reconstructions have prompted the use of DCRT's iPSC/860. This year progress has been made in combining many more images of our 3-D reconstruction, in part due to the availability of the Intel Supercomputer (see section on High Performance Biomedical Computing). Use of a new software program (EMPFTREF), provided by T.S. Baker (Purdue University) is providing substantially more accurate orientation refinement, and therefore much higher resolution 3-D reconstructions.
