Duane P. GrandgenettProfessor of Molecular Virology
Background: Ph. D. in Microbiology (University of Iowa, 1970)
The Retrovirus Integrase:
Numerous laboratories have made significant contributions towards understanding the mechanisms involved in retrovirus integration over the last 35 years. Integration of HIV-1 DNA into human chromosomes is essential for viral replication. Unchecked replication of HIV-1 in humans causes AIDS. Integration of the viral DNA genome is mediated by the viral integrase (IN). The FDA has approved several very effective IN strand transfer inhibitors which are used in combination with other inhibitors directed against the viral reverse transcriptase and protease to prevent AIDS.
Major Research Interests:
To investigate the structure-functional relationship between retrovirus IN and its viral DNA substrate in the assembled synaptic complex that mediates concerted integration.
Current Research Projects:
Following retrovirus infection of cells and subsequent reverse transcription of the viral RNA, linear viral DNA with IN and other viral proteins produce a cytoplasmic macromolecular structure termed the preintegration complex (PIC). The PIC is transported into the nucleus where the viral DNA ends are inserted in a concerted fashion by IN into the host chromosomes.
We are currently using purified recombinant HIV-1 and Rous sarcoma virus IN with viral DNA substrates to reconstruct the synaptic complex that mimics the ability of the PIC to promote concerted integration. Our efforts are to identify what IN protomer is used to assembly the IN tetramer that is necessary for the synaptic complex to catalyze concerted integration. Our published data suggests it is a monomer instead of a dimer for HIV-1 IN is the protomer for assembly. The assembly pathways for the synaptic complex using HIV-1 IN as well as Rous sarcoma virus IN are being investigated. Clinical strand transfer inhibitors of HIV-1 concerted integration are also being studied. Further crystallography studies on the RSV and HIV-1 synaptic complexes as well as these complexes in the present of target DNA molecules are being pursued.
Shi, K., Pandey, K. K., Bera, S., Vora, A., Grandgenett, D. P., and Aihara, H. (2013)
A possible role for the asymmetric C-terminal domain dimer of Rous sarcoma virus
integrase in viral DNA binding. Plos One 8:e56892.
Major Point: Crystal structure of the 3-domain Rous sarcoma virus IN was resolved at
1.86 Å. The binding of Rous sarcoma virus IN may be different than that observed with
the 4-domain prototype foamy virus IN to viral DNA.
Pandey, K. K., Bera, S., and Grandgenett, D. P. (2011) The HIV-1 integrase monomer
induces a specific interaction with LTR DNA for concerted integration. Biochemistry
Major Point: The HIV-1 IN monomer selectively interacts with the viral DNA ends
for concerted integration and appears to be the precursor of the IN tetramer
necessary for assembly of the synaptic complex. The monomer may be more
suitable than dimers of IN for producing crystals of IN/DNA complexes.
Bera, S., Pandey, K.K., Vora, A. and Grandgenett, D.P. (2011) HIV-1 integrase strand
transfer inhibitors stabilize an integrase-single blunt-ended DNA complex. J. Mol. Biol.
Major Point: Various strand transfer inhibitors at high concentrations promote the
formation of HIV-1 IN on a single viral DNA molecule.
Grandgenett, D. P. (2011). pp32 is Hot. In HIV-1 integrase: Mechanism and inhibitor design. ed., Neamati, N. and Wang, G. Wiley Press, June (Invited Book Chapter).
Major Point: A description of our research efforts that led to the discovery of the avian
retrovirus IN in 1978.
Grandgenett, D. .P, Korolev, S. (2010) Retrovirus Integrase-DNA structure elucidates
concerted integration mechanisms.Viruses. 2:1185-1189.
Major Point: Review of prototype foamy virus IN bound to viral DNA ends
in a crystal structure.
Pandey, K.K., Bera, S., Vora, A. C. and Grandgenett, D.P. (2010) Physically Trapping of the HIV-1 synaptic complex by different structural classes of integrase strand transfer inhibitors. Biochemistry 49:8376-8387.
Major Point: Clinical strand transfer inhibitor Raltegravir and others are able to “trap” or stabilize the HIV-1 synaptic complex by binding to the IN-DNA complex.
Bera, S., Pandey, K.K., Vora, A. C., and Grandgenett, D.P. (2009) Molecular interactions between HIV-1 integrase and the two viral DNA ends within the synaptic complex that mediates concerted integration. J. Mol. Biol. 389:183-198.
Major Point: The HIV-1 IN forms a specific complex on the viral DNA ends that produces an ~ 32 bp DNaseI protective footprint while protein-crosslinking of IN in the synaptic complex yields dimers and tetramers.
Grandgenett, D. P., Pandey, K. K., Bera, S., and Vora, A. C., Zahm, J., and Sinha, S. (2009). Biochemical and biophysical analyses of concerted (U3/U5) integration. In Mechanistic and Pharmacological Analyses of HIV-1 Integration, ed. Engelman, A., Elesvier, Methods 47:229-236.
Major Point: Description of our purification method for HIV-1 IN and concerted integration analysis. This technique is currently widely used to isolate the IN monomer which is utilized as a tool for evaluation of allosteric inhibitors, discovered in 2010 by others. These inhibitors inhibit LEDGF/p75 interactions with IN and also promote multerization of IN in virions thus inactivating the virus. Allosteric inhibitors of IN may have significant clinical application.
Pandey, K.K., Sinha, S., and Grandgenett, D. P. (2007). Transcriptional co-activator LEDGF/p75 modulates HIV-1 integrase mediated concerted integration. J. Virol. 81:3969-3979.
Major Point: We were first to demonstrate that the cellular cofactor LEDGF/p75 inhibits the concerted integration reaction if IN and LEDGF are incubated together first prior to addition of DNA. Modest stimulation of concerted integration is observed if the synaptic complex is formed prior to addition of LEDGF/p75.