Dennis Brown

My laboratory investigates the structure, function and assembly of the model membrane containing virus Sindbis. Sindbis virus is the prototype of the alpha viruses, a group of infectious agents which are transmitted in nature by blood-sucking insects. The alphaviruses are simple in composition (composed of multiple copies of three proteins) but complex in structure. The virus is made up of two structurally identical icosahedral protein shells between which is sandwiched a membrane bilayer. The two protein shells are connected to one another by protein-protein interactions which span the membrane bilayer. The protein associations result in a very strong but metastable structure capable of rapidly disassembling to release its genetic material (a single strand of RNA) into a host cell upon contacting an appropriate receptor.

The structure of the mature virion is being determined at high resolution using Electron Cryo-Microscopy. This technology uses massively parallel computers to analyze images of viruses obtained in high voltage electron microscopes equipped with liquid nitrogen stages and field emission guns. Data obtained by morphological studies are verified by biophysical and biochemical analysis of protein configuration and protein/protein associations in the virus structure. Our goal is to produce an image of the structure of this membrane containing virus at atomic resolution.

We are determining the pathway by which virus components are folded and associate with one another inside of the infected cell as the infectious virion is assembled from its component parts. We have determined that the two membrane glycoproteins which make up the outer icosahedral shell pass through a number of complex disulfide bridged intermediates as they are combined into the functional virion. The techniques of molecular and cellular biology together with molecular cloning and site-directed mutagenesis are employed to elucidate the pathway of virus assembly.