title: Unfolding the HIV-1 reverse transcriptase RNase H domain--how to lose a molecular tug-of-war.
abstract: Formation of the mature HIV-1 reverse transcriptase (RT) p66/p51 heterodimer requires subunit-specific processing of the p66/p66' homodimer precursor. Since the ribonuclease H (RH) domain contains an occult cleavage site located near its center, cleavage must occur either prior to folding or subsequent to unfolding. Recent NMR studies have identified a slow, subunit-specific RH domain unfolding process proposed to result from a residue tug-of-war between the polymerase and RH domains on the functionally inactive, p66' subunit. Here, we describe a structural comparison of the isolated RH domain with a domain swapped RH dimer that reveals several intrinsically destabilizing characteristics of the isolated domain that facilitate excursions of Tyr427 from its binding pocket and separation of helices B and D. These studies provide independent support for the subunit-selective RH domain unfolding pathway in which instability of the Tyr427 binding pocket facilitates its release followed by domain transfer, acting as a trigger for further RH domain destabilization and subsequent unfolding. As further support for this pathway, NMR studies demonstrate that addition of an RH active site-directed isoquinolone ligand retards the subunit-selective RH' domain unfolding behavior of the p66/p66' homodimer. This study demonstrates the feasibility of directly targeting RT maturation with therapeutics.
authors.affiliation: Genome Integrity and Structural Biology Laboratory, National Institute of Environmental health Sciences, NIH, Research Triangle Park, NC 27709, USA.
authors.forename: Xunhai
authors.initials: X
authors.lastname: Zheng
authors.affiliation: Genome Integrity and Structural Biology Laboratory, National Institute of Environmental health Sciences, NIH, Research Triangle Park, NC 27709, USA.
authors.forename: Lars C
authors.initials: LC
authors.lastname: Pedersen
authors.affiliation: Genome Integrity and Structural Biology Laboratory, National Institute of Environmental health Sciences, NIH, Research Triangle Park, NC 27709, USA.
authors.forename: Scott A
authors.initials: SA
authors.lastname: Gabel
authors.affiliation: Genome Integrity and Structural Biology Laboratory, National Institute of Environmental health Sciences, NIH, Research Triangle Park, NC 27709, USA.
authors.forename: Geoffrey A
authors.initials: GA
authors.lastname: Mueller
authors.affiliation: Genome Integrity and Structural Biology Laboratory, National Institute of Environmental health Sciences, NIH, Research Triangle Park, NC 27709, USA.
authors.forename: Eugene F
authors.initials: EF
authors.lastname: DeRose
authors.affiliation: Genome Integrity and Structural Biology Laboratory, National Institute of Environmental health Sciences, NIH, Research Triangle Park, NC 27709, USA london@niehs.nih.gov.
authors.forename: Robert E
authors.initials: RE
authors.lastname: London
mesh_terms: D001665:Binding Sites; D020134:Catalytic Domain; D019281:Dimerization; D054303:HIV Reverse Transcriptase / Q000737:chemistry / Q000235:genetics*; D015497:HIV-1 / Q000201:enzymology* / Q000235:genetics / Q000472:pathogenicity; D006801:Humans; D011487:Protein Conformation; D017434:Protein Structure, Tertiary / Q000235:genetics; D018894:Reverse Transcriptase Inhibitors / Q000008:administration & dosage; D016914:Ribonuclease H / Q000737:chemistry / Q000235:genetics
chemical_list: D018894:Reverse Transcriptase Inhibitors; C514824:reverse transcriptase, Human immunodeficiency virus 1; D054303:HIV Reverse Transcriptase; D016914:Ribonuclease H