The goal of this project is to crystallize and determine the X-ray structure of the Vpu channel protein of the human immunodeficiency virus type-1 (HIV-1). The insights into Vpu function provided by this structure should contribute materially to the development of novel AIDS therapies. This work is part of a long term effort to elucidate the structures of membrane-bound proteins and to probe the functional significance of these structures. Vpu is an integral membrane protein with channel activity that mediates two important function in the life of HIV-1: It controls budding and release of new virions from the surface of the infected cell, and it induces the destruction of the host's CD4 protein within the cell. After infection of cells in vitro, isolates of HIV-1 lacking Vpu release far fewer virus particles into the medium than wild type virus, supporting the notion that the action of this protein is important to virus action. In addition to its importance as a potential AIDS drug target, at 81 amino acids, Vpu is an excellent model system for the development of new membrane protein structural methods.
The specific aims of this project are devoted to a rational strategy for obtaining crystals of detergent-solubilized Vpu, and to using these crystals to elucidate the three-dimensional structure of the protein. A vesicle-based assay for functional Vpu channel will be developed and used to estimate the subunit stoichiometry of the channel. Detergents will be screened for their ability to maintain this stoichiometry in solution. Appropriate detergents will then be used to develop crystallization assays designed to simultaneously titrate the solubilities of the protein and detergent components of the Vpu-detergent complex. Promising conditions will be optimized by statistically-based methods, and the crystals obtained will be used to determine the X-ray crystal structure of the protein.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Program Projects (P01)
Project #
5P01GM056538-02
Application #
6107810
Study Section
Project Start
1998-08-01
Project End
1999-07-31
Budget Start
1997-10-01
Budget End
1998-09-30
Support Year
2
Fiscal Year
1998
Total Cost
Indirect Cost
Name
University of Pennsylvania
Department
Type
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
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Mesleh, Michael F; Opella, Stanley J (2003) Dipolar Waves as NMR maps of helices in proteins. J Magn Reson 163:288-99

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