title: Structures of HIV-1 RT-RNA/DNA ternary complexes with dATP and nevirapine reveal conformational flexibility of RNA/DNA: insights into requirements for RNase H cleavage.
abstract: In synthesizing a double-stranded DNA from viral RNA, HIV-1 reverse transcriptase (RT) generates an RNA/DNA intermediate. RT also degrades the RNA strand and synthesizes the second DNA strand. The RNase H active site of RT functions as a nuclease to cleave the RNA strand; however, the structural basis for endonucleolytic cleavage of the RNA strand remains elusive. Here we report crystal structures of RT-RNA/DNA-dATP and RT-RNA/DNA-nevirapine (NVP) ternary complexes at 2.5 and 2.9 Å resolution, respectively. The polymerase region of RT-RNA/DNA-dATP complex resembles DNA/DNA ternary complexes apart from additional interactions of 2'-OH groups of the RNA strand. The conformation and binding of RNA/DNA deviates significantly after the seventh nucleotide versus a DNA/DNA substrate. Binding of NVP slides the RNA/DNA non-uniformly over RT, and the RNA strand moves closer to the RNase H active site. Two additional structures, one containing a gapped RNA and another a bulged RNA, reveal that conformational changes of an RNA/DNA and increased interactions with the RNase H domain, including the interaction of a 2'-OH with N474, help to position the RNA nearer to the active site. The structures and existing biochemical data suggest a nucleic acid conformation-induced mechanism for guiding cleavage of the RNA strand.
authors.affiliation: Center for Advanced Biotechnology and Medicine, Department of Chemistry and Chemical Biology, Rutgers University, Piscataway, NJ 08854, USA.
authors.forename: Kalyan
authors.initials: K
authors.lastname: Das
authors.affiliation: Center for Advanced Biotechnology and Medicine, Department of Chemistry and Chemical Biology, Rutgers University, Piscataway, NJ 08854, USA.
authors.forename: Sergio E
authors.initials: SE
authors.lastname: Martinez
authors.affiliation: Center for Advanced Biotechnology and Medicine, Department of Chemistry and Chemical Biology, Rutgers University, Piscataway, NJ 08854, USA.
authors.forename: Rajiv P
authors.initials: RP
authors.lastname: Bandwar
authors.affiliation: Center for Advanced Biotechnology and Medicine, Department of Chemistry and Chemical Biology, Rutgers University, Piscataway, NJ 08854, USA arnold@cabm.rutgers.edu.
authors.forename: Eddy
authors.initials: E
authors.lastname: Arnold
mesh_terms: D019380:Anti-HIV Agents / Q000737:chemistry*; D004279:DNA, Viral / Q000737:chemistry* / Q000378:metabolism; D003838:Deoxyadenine Nucleotides / Q000737:chemistry*; D054303:HIV Reverse Transcriptase / Q000737:chemistry*; D008958:Models, Molecular; D019829:Nevirapine / Q000737:chemistry*; D009690:Nucleic Acid Conformation; D012367:RNA, Viral / Q000737:chemistry* / Q000378:metabolism; D018894:Reverse Transcriptase Inhibitors / Q000737:chemistry*; D016914:Ribonuclease H / Q000378:metabolism
chemical_list: D019380:Anti-HIV Agents; D004279:DNA, Viral; D003838:Deoxyadenine Nucleotides; D012367:RNA, Viral; D018894:Reverse Transcriptase Inhibitors; D019829:Nevirapine; C514824:reverse transcriptase, Human immunodeficiency virus 1; D054303:HIV Reverse Transcriptase; D016914:Ribonuclease H; C026600:2'-deoxyadenosine triphosphate