A Homology Model of HIV-1 Integrase and Analysis of Mutations Designed to Test the Model

Johnson, Barry C.; Métifiot, Mathieu; Ferris, Andrea L.; Pommier, ﻿Yves; Hughes, Stephen H.

doi:10.1016/j.jmb.2013.03.027

Cited by 47 publications

(61 citation statements)

References 51 publications

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“…While no crystal structure for the intact HIV-1 IN currently exists, as either part of the GagPol precursor or part of the intasome or any other state, we used recently published models of the HIV-1 intasome to depict our IN mutations in the context of a plausible IN structure (Fig. 4A) (28,29). The two published models are similar, and our analyses below do not depend on which model is actually used.…”

Section: Resultsmentioning

confidence: 99%

“…Examination of the HIV-1 intasome was done with structure coordinates provided by Stephen Hughes (28) and Alan Engelman (29) using MacPyMOL. UCSF Chimera was utilized for analysis of solvent-accessible surface area.…”

Section: Plasmids and Constructionmentioning

confidence: 99%

“…Determination of the crystal structure of the prototype foamy virus (PFV) intasome illuminated how IN domains contribute to protein multimerization, as well as to vDNA and host DNA binding (26,27). By analogy to the PFV intasome crystal structure, new models of the HIV-1 intasome structure have been derived (28,29).…”

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confidence: 99%

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Uneven Genetic Robustness of HIV-1 Integrase

Rihn

Hughes

Wilson

et al. 2015

J Virol

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Genetic robustness (tolerance of mutation) may be a naturally selected property in some viruses, because it should enhance adaptability. Robustness should be especially beneficial to viruses like HIV-1 that exhibit high mutation rates and exist in immunologically hostile environments. Surprisingly, however, the HIV-1 capsid protein (CA) exhibits extreme fragility. To determine whether fragility is a general property of HIV-1 proteins, we created a large library of random, single-amino-acid mutants in HIV-1 integrase (IN), covering >40% of amino acid positions. Despite similar degrees of sequence variation in naturally occurring IN and CA sequences, we found that HIV-1 IN was significantly more robust than CA, with random nonsilent IN mutations only half as likely to cause lethal defects. Interestingly, IN and CA were similar in that a subset of mutations with high in vitro fitness were rare in natural populations. IN mutations of this type were more likely to occur in the buried interior of the modeled HIV-1 intasome, suggesting that even very subtle fitness effects suppress variation in natural HIV-1 populations. Lethal mutations, in particular those that perturbed particle production, proteolytic processing, and particle-associated IN levels, were strikingly localized at specific IN subunit interfaces. This observation strongly suggests that binding interactions between particular IN subunits regulate proteolysis during HIV-1 virion morphogenesis. Overall, use of the IN mutant library in conjunction with structural models demonstrates the overall robustness of IN and highlights particular regions of vulnerability that may be targeted in therapeutic interventions. IMPORTANCEThe HIV-1 integrase (IN) protein is responsible for the integration of the viral genome into the host cell chromosome. To measure the capacity of IN to maintain function in the face of mutation, and to probe structure/function relationships, we created a library of random single-amino-acid IN mutations that could mimic the types of mutations that naturally occur during HIV-1 infection. Previously, we measured the robustness of HIV-1 capsid in this manner and determined that it is extremely intolerant of mutation. In contrast to CA, HIV-1 IN proved relatively robust, with far fewer mutations causing lethal defects. However, when we subsequently mapped the lethal mutations onto a model of the structure of the multisubunit IN-viral DNA complex, we found the lethal mutations that caused virus morphogenesis defects tended to be highly localized at subunit interfaces. This discovery of vulnerable regions of HIV-1 IN could inform development of novel therapeutics.

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Section: Resultsmentioning

confidence: 99%

Section: Plasmids and Constructionmentioning

confidence: 99%

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Uneven Genetic Robustness of HIV-1 Integrase

Rihn

Hughes

Wilson

et al. 2015

J Virol

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“…In the NMR structure of the isolated CTD, which is seen as a dimer with Lys-264 and Lys-266 being fully surface-exposed and positioned on the opposite side of the CTD-CTD interface (28 -30). In the HIV-1 intasome models, which have been generated based on the analogous prototype foamy virus intasome structure, the CTD domains do not engage in protein-protein interactions and instead interact with viral DNA (10,54,55). In particular, Lys-264 and Lys-266 directly contact viral DNA (Fig.…”

Section: Discussionmentioning

confidence: 99%

A Critical Role of the C-terminal Segment for Allosteric Inhibitor-induced Aberrant Multimerization of HIV-1 Integrase

Shkriabai

Dharmarajan

Slaughter

et al. 2014

Journal of Biological Chemistry

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Background: Allosteric integrase (IN) inhibitors (ALLINIs) promote aberrant protein multimerization. Results: ALLINI-2 induces protein-protein interactions, including C-terminal residues Lys-264 and Lys-266, which lead to aberrant, higher-order IN multimerization. Conclusion: The protein-protein contacts beyond the inhibitor binding site contribute to aberrant IN multimerization. Significance: Our findings provide structural clues and underscore the significance for exploiting IN multimerization as a new therapeutic target.

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“…2A) or ASLV (77), will support the formation of the dimer-of-dimers architecture observed with the PFV intasome. Numerous groups have assembled HIV-1 models based on the PFV structure (105,135,192,193), indicating reasonable potential for concordance (64). With ASLV integrase NTD-CCD and CCD-CTD linkers as seemingly short as 13 and 8 amino acid residues, respectively (77), the structure of the ASLV intasome should shed light on the potential universality of the PFV intasome architecture as a virus family-wide model.…”

Section: Future Directions In Integrase Structural Biologymentioning

confidence: 99%

Retroviral Integrase Structure and DNA Recombination Mechanism

Engelman

Cherepanov

2014

Microbiol Spectr

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SUMMARYDue to the importance of human immunodeficiency virus type 1 (HIV-1) integrase as a drug target, the biochemistry and structural aspects of retroviral DNA integration have been the focus of intensive research during the past three decades. The retroviral integrase enzyme acts on the linear double-stranded viral DNA product of reverse transcription. Integrase cleaves specific phosphodiester bonds near the viral DNA ends during the 3′ processing reaction. The enzyme then uses the resulting viral DNA 3′-OH groups during strand transfer to cut chromosomal target DNA, which simultaneously joins both viral DNA ends to target DNA 5′-phosphates. Both reactions proceed via direct transesterification of scissile phosphodiester bonds by attacking nucleophiles: a water molecule for 3′ processing, and the viral DNA 3′-OH for strand transfer. X-ray crystal structures of prototype foamy virus integrase-DNA complexes revealed the architectures of the key nucleoprotein complexes that form sequentially during the integration process and explained the roles of active site metal ions in catalysis. X-ray crystallography furthermore elucidated the mechanism of action of HIV-1 integrase strand transfer inhibitors, which are currently used to treat AIDS patients, and provided valuable insights into the mechanisms of viral drug resistance.

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A Homology Model of HIV-1 Integrase and Analysis of Mutations Designed to Test the Model

Cited by 47 publications

References 51 publications

Uneven Genetic Robustness of HIV-1 Integrase

Uneven Genetic Robustness of HIV-1 Integrase

A Critical Role of the C-terminal Segment for Allosteric Inhibitor-induced Aberrant Multimerization of HIV-1 Integrase

Retroviral Integrase Structure and DNA Recombination Mechanism

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