Discipline: Biological Sciences
Subcategory: Biochemistry (not Cell and Molecular Biology and Genetics)
Room: Exhibit Hall A
Gilberto Perez - University of Missouri - Columbia
Co-Author(s): Rohit Rao, University of Missouri, Columbia, MO Anna Davis, University of Missouri, Columbia, MO Kyle J. Hill, University of Missouri, Columbia, MO Anders Sönnerborg, Karokinska Institutet, Stockholm, Sweden Ujjwal Neogi, Karokinska Institutet, Stockholm, Sweden Kamlendra Singh, University of Missouri, Columbia, MO
Currently recommended anti-HIV treatment includes one HIV-1 integrase (HIV-1 IN) inhibitor (INIs) in the backbone of two nucleoside reverse transcriptase inhibitors (NRTIs). Integrase (IN), 32 kDa protein is encoded by the pol gene of HIV-1. The role of this enzyme is to integrate reverse transcribed double stranded DNA (dsDNA) from the RNA gene of HIV-1 into the host chromosome. To integrate this dsDNA into the host genome, IN conducts two reactions: (i) the 3’ end processing or 3’-EP reaction, which is the cleavage of a dinucleotide from both 3’ ends of viral DNA and (ii) the strand transfer (ST) reaction, which is inserting of viral DNA with exposed 3’OH groups and covalently attaching it to the host genome. Despite, a higher efficacy of INIs, the virus emerges with the resistance mutations against these drugs. In addition, subtype-specific polymorphisms (PMs) can affect the outcome of INI-based therapy. Here we report the biochemical characterization of an IN derived from a patient infected with CRF01_AE subtype. The IN from this strain contained M50I polymorphism. The DNA binding determined by Microscale Thermophoresis (MST) assays showed that CRF01_AE IN binds DNA with an affinity of 24 nM, which is marginally better than that of that of IN derived from the laboratory strain (HXB2) (32 nM). The increased DNA binding activity of CRF01_AE IN resulted in an increase in its 3’-EP activity and showed that this IN excises the dinucleotide from 3’-end with a greater efficiency compared to HXB2-derived IN. Since, there needs to be a balance between inhibition and catalysis, our results show that INIs may not be an effective drug against CRF01_AE IN containing M50I polymorphism which affects drug susceptibility.
Funder Acknowledgement(s): NIH IMSD Grant # R25GM056901
Faculty Advisor: Dr. Kamlendra Singh, SinghKa@mail.missouri.edu
Role: Purification of Protein, Miscroscale Thermophoresis Assays, and Gel Assays.Helped with data analysis.