Can anyone explain to me what this paper means??? - Best, Rivka _______ http://antiviralresistance.org/abstr...ster5_2010.pdf POSTER 5 BIOCHEMICAL, STRUCTURAL, AND INHIBITION STUDIES OF XMRV REVERSE TRANSCRIPTASE Tanya Ndognwe1, Karen Kirby1, Bruno Marchand1, Adeyemi Adedeji1, Eleftherios Michailidis1, Yee-Tsuei Ong1, Atsuko Hachiya1, Emily Ryan1, Shun-Lu Liu1, Angela Whatley1, Donald H. Burke1, Sanath Kumar1, Marc Johnson1, Ei-Ichi Kodama2, Krista A. Delviks-Frankenberry3, Vinay K. Pathak3, Hiroaki Mitsuya4, Michael A. Parniak5, Kamal Singh1, and Stefan G. Sarafianos1 1Department of Molecular Microbiology & Immunology, University of Missouri School of Medicine, Columbia, MO; 2Division of Emerging Infectious Diseases, Tohoku University School of Medicine, Sendai, Japan; 3HIV Drug Resistance Program, National Cancer Institute, Frederick, MD; 4Department of Internal Medicine, Kumamoto University School of Medicine, Kumamoto, Japan & Experimental Retrovirology Section, HIV/AIDS Malignancy Branch, NIH, Bethesda, MD; 5Department of Molecular Genetics & Biochemistry, University of Pittsburgh School of Medicine, Pittsburgh, PA The Xenotropic Murine Leukemia Virus-Related Virus (XMRV) was originally identified in biological samples from familial prostate cancer patients and was more recently reported in patients with chronic fatigue syndrome (CFS). Although other studies have failed to detect XMRV in CFS or prostate cancer patients, given the potential importance of this virus for human disease we initiated studies on its replication mechanism and susceptibility to inhibitors. We used biochemical, biophysical, virological, and structural methods to characterize the DNA polymerase and RNase H functions of XMRV reverse transcriptase (RT). We compared the properties of XMRV RT, XMRV RNase H, Moloney Murine Leukemia Virus (MuLV) RT, and HIV RT. Using steady state and pre-steady state kinetics we demonstrated that XMRV RT is the slowest and least efficient of the three enzymes in synthesizing DNA or cleaving RNA/DNA. Similarly, its ability to unblock chain-terminated primers using PPi or ATP are much lower compared to HIV RT. Its reduced polymerase and RNAse H activity is due in part to a lower affinity for nucleic acid, as judged by gel-shift assays and confirmed by surface plasmon resonance experiments, which revealed that the deficiency in DNA binding is due to a high dissociation rate. Trap experiments showed that XMRV RT has very low processivity compared to HIV RT. Transient kinetics of mismatch incorporations revealed that XMRV RT has higher fidelity than MuLV and HIV RTs. Nonetheless, XMRV and MuLV appear to have comparable fidelities in cell-based assays. The polymerase function of XMRV RT is susceptible to antiretrovirals from several classes, but not to NNRTIs. XMRV RT is inhibited efficiently by AZT-TP, PMEADP, d4T-TP, PMPA-DP, and by a nucleic acid aptamer. Two potent NRTIs that block XMRV RT efficiently by a different mechanism also have potent antiviral activity in pseudotype-based and replication competent virus-based assays. We have also identified compounds that efficiently block the RNase H activity of XMRV RT and of active XMRV RNase H fragments. Finally, we have solved the crystal structure of XMRV RNase H at high resolution (1.5 ), which will facilitate the design of new and more potent XMRV inhibitors.