HIV Interaction and Viral Evolution Center Research
Research Highlights
The central theme of the HIVE Center is to further our structural and functional knowledge of HIV assembly, maturation, reverse transcription, and integration with the human host with the goal of determining the atomic resolution structures and the dynamic mechanisms of the macromolecular complexes involved in these processes. The ultimate aim is to relate this knowledge to the evolution of drug resistance and to the improvement of drug design methodologies in the treatment of HIV infected individuals.
2013 Research Highlights
Allosteric Integrase Inhibitor Potency is Determined Through the Inhibition of HIV-1 Particle Maturation
Allosteric integrase inhibitors, originally developed to block viral integration, are also surprisingly found to be potent inhibitors of viral maturation.
The A128T Resistance Mutation Reveals Aberrant ProteinMultimerization as the Primary Mechanism of Action of Allosteric HIV-1Integrase Inhibitors.
PubMed Central PMCID: PMC3668738 (embargoed until May 31 2104)
Structural basis for high-affinity binding of LEDGF PWWP to mononucleosomes.
This study reveals that cellular LEDGF/p75 is tightly bound to mononucleosomes, and cooperative binding of the hydrophobic cavity and basic surface to the cognate histone peptide and DNA wrapped in mononucleosomes is essential for high-affinity binding to chromatin.
Eidahl JO, Crowe BL, North JA, McKee CJ, Shkriabai N, Feng L, Plumb M, Graham RL, Gorelick RJ, Hess S, Poirier MG, Foster MP, Kvaratskhelia M. Nucleic Acids Res. 2013 Apr 1;41(6):3924-36.
Snapshot of the equilibrium dynamics of a drug bound to HIV-1 reverse transcriptase.
Femtosecond experiments and theory expose the molecular level dynamics of rilpivirine bound to HIV-1 RT.
Detecting allosteric sites of HIV-1 reverse transcriptase by X-ray crystallographic fragment screening
New allosteric sites for inhibitor binding were discovered in HIV reverse transcriptase, providing opportunities for discovery of novel anti-HIV drugs.
Rapid deep sequencing of patient-derived HIV with ion semiconductor technology
Ion Torrent devices are used for deep sequencing of drug resistant HIV samples, yielding high levels of sequencing coverage in HIV Gag and protease, and allowing the detection of mutations at low frequencies.
Chang MW, Oliveira G, Yuan J, Okulicz JF, Levy S, Torbett BE. J Virol Methods. 189(1):232-4 (2013).
Allosteric Inhibition of HIV-1 Integrase Activity
A review of progress with the development of allosteric integrase inhibitors (ALLINIs) that compete with LEDGF/p75 for binding to integrase, disrupting viral maturation and inhibiting integrase function.
Engelman, A., Kessl, J. J. & Kvaratskhelia, M. Curr. Op. Chem. Biol. 17, 339-345 (2013)
PubMed Central PMCID: PMC3679204 (embargoed until June 1 2014)
Small Molecule Regulation of Protein Conformation by Binding in the Flap of HIV Protease
Crystallographic structures of two small indoles reveal a binding site that favors a closed conformation of the HIV protease flaps.
Preliminary Work
Structural and Functional Insights into Alphavirus Polyprotein Processing and PathogenesisThe alphavirus replication machinery consists of four nonstructural proteins produced as a single polyprotein. The structure has been solved of P23 in a precleavage form. The P2/3 cleavage site is located at the base of a narrow cleft and is not readily accessible, and the nsP2 protease active site is over 40 Angstroms away, supporting a regulated, trans cleavage mechanism.
G. Shin, S. A. Yost, M. T. Miller, E. J. Elrod, A. Grakoui & J. Marcotrigiano (2012) Proc. Natl. Acad. Sci. USA 109, 16534-16539.
A comparison of the ability of rilpivirine (TMC278) and selected analogues to inhibit clinically relevant HIV-1 reverse transcriptase mutants.
A combination of structure activity relationships and X-ray crystallography was used to examine non-nucleoside reverse transcriptase inhibitors that are structurally related to rilpivirine to determine their ability to inhibit wildtype reverse transcriptase and several clinically relevant mutants.
HIV-1 Reverse Transcriptase Complex with DNA and Nevirapine Reveals Non-nucleoside Inhibition Mechanism
The crystal structures of reverse transcriptase with DNA and two types of inhibitors have been solved. The RT-DNA complex in the crystal could bind either the non-nucleoside inhibitor nevirapine or the nucleoside inhibitor AST-triphosphate, but not both. The structures reveal the complementary roles these different classes of inhibitor play in widely-used anti-AIDS therapies.
K. Das, S. E. Martinez & E. Arnold (2012) Nat. Struct. Mol. Biol. 19, 253-259.
Retroviral Intasome Assembly and Inhibition of DNA Strand Transfer
The structure was solved of full-length retroviral integrase from prototype foamy virus in complex with its cognate DNA. The structure shows the organization of retroviral intasome , with an integrase tetramer tightly associated with a pair of viral DNA ends. The structure reveals the extensive protein-DNA and protein-protein interactions involved in retroviral integration, and provides a model for the HIV-1 intasome.
S. Hare, S. S. Gupta, E. Valkov, A. Engelman & P. Cherepanov (2010) Nature 464, 232-236.
Fragment-Based Screen Against HIV Protease
Two new inhibitor binding sites were discovered on wild-type HIV protease using fragment-based screening techniques. Protease was cocrystallized or soaked with chemical fragments using five different crystal forms, and 378 data sets were collected. Fragment binding induces a new conformation and crystal form in protease with the active site inhibitor TL-3. This study is the first fragment-based crystallographic screen against HIV protease, revealing two new exosites that stabilize inhibitor binding at the active site.
A. L. Perryman, Q. Zhang, H. H. Soutter, R. Rosenfeld, D. E. McRee, A. J. Olson, J. E. Elder & C. D. Stout (2010) Chem Biol. Drug Des. 75, 257-268.
Theory of binless multi-state free energy estimation with applications to protein-ligand binding.
The paper describes a simplified technique for computing free energies and expectations from multiple ensembles.
Tan Z, Gallicchio E, Lapelosa M, Levy RM. J Chem Phys. 136(14):144102 (2012).
