We propose X-ray crystallographic structural studies of the envelope glycoproteins from HIV, the virus that cause AIDS. A long-range goal is to use structural information about receptor- binding, fusion activity and antigenicity to design inhibitors and to improve potential vaccines. The project requires definition of stable, well-folded species for rational crystallization efforts. Important information about the properties of the gp120-gp41 complex will be obtained in such experiments. We will begin with secreted gp120 and membrane-bound gp160 from genetically engineered mammalian and insect-cell vectors. We will seek to prepare a soluble (i.e., membrane-anchorless) form of gp160 or of gp120-gp41 by genetic deletion and by limited proteolysis. We will examine glycoproteins derived from several isolates of HIV, and we will explore the effect of carbohydrate content by using glycosidase-treated and genetically-engineered carbohydrate deficient proteins. We will search for a minimum T4 binding domain, using limited proteolysis and end-deletion mutagenesis of secreted gp120, since preparation of such a domain might facilitate study of virus-receptor interaction, including crystallization of a complex with soluble T4. We will determine the oligomeric state, proteolytic susceptibility, T4 binding, antibody binding, and membrane-fusion activity of all forms of the glycoprotein to be crystallized, and we will compare these properties with those of native glycoprotein (gp120-gp41) from stably infected, non-virus-producing cells.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI025287-03
Application #
3138706
Study Section
(SRC)
Project Start
1987-08-01
Project End
1992-07-31
Budget Start
1989-08-01
Budget End
1990-07-31
Support Year
3
Fiscal Year
1989
Total Cost
Indirect Cost
Name
Harvard University
Department
Type
Schools of Arts and Sciences
DUNS #
071723621
City
Cambridge
State
MA
Country
United States
Zip Code
02138
Schawaller, M; Smith, G E; Skehel, J J et al. (1989) Studies with crosslinking reagents on the oligomeric structure of the env glycoprotein of HIV. Virology 172:367-9