The X-ray structures of the 4-domain prototype foamy virus (PFV) integrase (IN)(1,2) and very recently the 3-domain Rous sarcoma virus (RSV) IN (3) in complex with viral DNA and/or viral-target DNA substrates have been determined. Yet, the crystal structure of the medically important 3- domain HIV IN by itself or in complex with DNA is still elusive. We propose to provide the means and methods using monomeric wild type (wt) HIV IN or modified IN constructs to accomplish this very difficult but highly significant project. The first necessary step to achieve this important goal is harvesting the ability of HIV IN to assemble these IN-DNA complexes at high protein concentrations in the ~100 to 150 M range for crystal screening and other in-solution structural investigations. Our recent success in defining new assembly pathways for stabilizing 3-domain IN-DNA complexes and the use of newly enhanced solubility methods for recombinant HIV IN will advance our investigative studies.

Public Health Relevance

It is necessary to obtain the 3-dimensional structure of the HIV IN in complex with viral DNA which currently does not exist to better understand the mechanism of integrase inhibitor interactions and drug resistance. We propose to facilitate the isolation of kinetically stabilized HIV IN-DNA complexes at high concentrations in-solution and screening of these complexes for crystal formation. With resolved IN-DNA structures, new allosteric inhibitor sites outside the active catalytic site in the presence of DNA may be identified and possibly provide mechanisms for pre-prophylaxis treatment.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Exploratory/Developmental Grants (R21)
Project #
5R21AI127196-02
Application #
9294971
Study Section
AIDS Molecular and Cellular Biology Study Section (AMCB)
Program Officer
Kuo, Lillian S
Project Start
2016-06-15
Project End
2018-05-31
Budget Start
2017-06-01
Budget End
2018-05-31
Support Year
2
Fiscal Year
2017
Total Cost
$227,250
Indirect Cost
$77,250
Name
Saint Louis University
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
050220722
City
Saint Louis
State
MO
Country
United States
Zip Code
63103