Pre-organized structure of viral DNA at the binding-processing site of HIV-1 integrase

Renisio, Jean-Guillaume

doi:10.1093/nar/gki346

Cited by 23 publications

(21 citation statements)

References 75 publications

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“…5, E and F) in agreement with the previous observation of DNA binding in the absence and presence of metal (26,27). The recently reported flexibility of the viral DNA end may allow cross-linking of nonprocessed substrate (28). Together, these experiments indicate that the integrase residue 148 is in the vicinity of the cytosine of the 5Ј-overhang and that catalysis is not required for DNA cross-linking.…”

Section: Speciessupporting

confidence: 89%

Integration Requires a Specific Interaction of the Donor DNA Terminal 5′-Cytosine with Glutamine 148 of the HIV-1 Integrase Flexible Loop

Johnson¹,

Santos

Pais

et al. 2006

Journal of Biological Chemistry

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Integration is essential for retroviral replication and gene therapy using retroviral vectors. Human immunodeficiency virus, type 1 (HIV-1), integrase specifically recognizes the terminal sequences of each long terminal repeat (LTR) and cleaves the 3-end terminal dinucleotide 5-GT. The exposed 3-hydroxyl is then positioned for nucleophilic attack and subsequent strand transfer into another DNA duplex (target or chromosomal DNA). We report that both the terminal cytosine at the protruding 5-end of the long terminal repeats (5-C) and the integrase residue Gln-148 are critical for strand transfer. Proximity of the 5-C and Gln-148 was demonstrated by disulfide cross-linking. Cross-linking is inhibited by the inhibitor 5CITEP 1-(5-chloroindol-3-yl)-3-hydroxy-3-(2H-tetrazol-5-yl)-propenone. We propose that strand transfer requires a conformational change of the integrase-viral (donor) DNA complex with formation of an H-bond between the N-3 of the 5-C and the amine group of Gln-148. These findings have implications for the molecular mechanisms coupling 3-processing and strand transfer as well as for the molecular pharmacology of integrase inhibitors.

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Section: Speciessupporting

confidence: 89%

Integration Requires a Specific Interaction of the Donor DNA Terminal 5′-Cytosine with Glutamine 148 of the HIV-1 Integrase Flexible Loop

Johnson¹,

Santos

Pais

et al. 2006

Journal of Biological Chemistry

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“…We therefore suggest that DAP prevents the local A/T pair destabilization that is essential for efficient processing. The necessity of this destabilization also found for avian sarcoma virus integrase (42) may account for the strict conservation of the CA/TG sequence in integrase substrates, as intrinsic conformational mobility is known to favor DNA flexibility (43)(44)(45)(46)(47). Such a local destabilization allows IN activity, even at an internal position, as seen in the internal processing of a 26-mer duplex (26B/A).…”

Section: Discussionmentioning

confidence: 90%

“…The DNA is locally distorted from a regular B form to a highly strained kinked conformation, leading to unwinding of the DNA, unstacking of the two central base pairs of the recognition site, and bending of the DNA (50). For IN, this bending and unstacking may be favored by the conformational mobility of the CA/TG fragment (43-44) and a loss of regular base stacking (45).…”

Section: Discussionmentioning

confidence: 99%

Probing of HIV-1 Integrase/DNA Interactions Using Novel Analogs of Viral DNA

Agapkina

Smolov

Barbe

et al. 2006

Journal of Biological Chemistry

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The specific activity of the human immunodeficiency virus, type 1 (HIV-1), integrase on the viral long terminal repeat requires the binding of the enzyme to certain sequences located in the U3 and U5 regions at the ends of viral DNA, but the determinants of this specific DNA-protein recognition are not yet completely understood. We synthesized DNA duplexes mimicking the U5 region and containing either 2-modified nucleosides or 1,3-propanediol insertions and studied their interactions with HIV-1 integrase, using Mn 2؉ or Mg 2؉ ions as integrase cofactors. These DNA modifications had no strong effect on integrase binding to the substrate analogs but significantly affected 3-end processing rate. The effects of nucleoside modifications at positions 5, 6, and especially 3 strongly depended on the cationic cofactor used. These effects were much more pronounced in the presence of Mg 2؉ than in the presence of Mn 2؉ . Modifications of base pairs 7-9 affected 3-end processing equally in the presence of both ions. Adenine from the 3rd bp is thought to form at least two hydrogen bonds with integrase that are crucial for specific DNA recognition. The complementary base, thymine, is not important for integrase activity. For other positions, our results suggest that integrase recognizes a fine structure of the sugar-phosphate backbone rather than heterocyclic bases. Integrase interactions with the unprocessed strand at positions 5-8 are more important than interactions with the processed strand for specific substrate recognition. Based on our results, we suggest a model for integrase interaction with the U5 substrate.Following reverse transcription, a DNA copy of the human immunodeficiency virus, type 1 (HIV-1), 2 RNA is integrated into the genome of infected cells. Integration is a prerequisite for viral replication and is catalyzed by the viral enzyme integrase (IN). IN binds to sequences located at the end of U3 and U5 parts of long terminal repeats (LTRs) of viral DNA and catalyzes the trimming, or 3Ј-end processing, of the terminal dinucleotide from the 3Ј-ends of both strands of the DNA. IN then mediates a strand transfer reaction that inserts the viral DNA into the host DNA. During this reaction, IN must bind simultaneously to viral and target DNA. However, IN interacts with these two DNA molecules in different ways as follows: binding to host DNA does not depend directly on host DNA sequence, whereas interaction with the viral DNA is a sequence-specific process. Nevertheless, the U5 and U3 sequences recognized by IN are not exactly identical.Strand transfer and 3Ј-end processing reactions may be carried out in vitro, using recombinant HIV IN, DNA duplexes mimicking U3 or U5 sequences of LTRs, and divalent metal ions, such as Mg 2ϩ or Mn 2ϩ . However, the Mn 2ϩ -and Mg 2ϩ -dependent activities of IN are not equivalent, with lower specificity reported for Mn 2ϩ -dependent IN (1, 2). Moreover, the inhibition of HIV-1 IN by compounds such as ␤-diketo acids, which interact with the active site of HIV-1 IN, is also metal-de...

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“…Their non-globular and elongated shapes prevent an overall structure determination based only on local relative restraints, while the property of RDCs to be referred to a unique molecular frame gives access to the long-range information needed to explore their overall shape. Nevertheless, few direct applications of RDC structure determinations of DNA 26,46,47 currently exist, whereas for RNA it is well established that RDCs are powerful agents for resolving their structures in solution. 48 -51 The present work based on a non-labelled DNA duplex confirms the improvement allowed by a restricted set of RDCs.…”

Section: Discussionmentioning

confidence: 99%

NMR assessment of the global shape of a non‐labelled DNA dodecamer containing a tandem of GT mismatches

Alvarez-Salgado

Desvaux

Boulard

2006

Magnetic Reson in Chemistry

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We have carried out a solution study of two non-labelled self-complementary DNA dodecamers d(GACTGTACAGTC)2 and d(GACTGTGCAGTC)2 by NMR, the second sequence composed of two G-T mismatches. Structures were determined using distances extracted from NOE effects alone or using both NOE and RDC constraints, measured in three different liquid crystalline media. We ensured that our data on the influence of the mesogen on the DNA structures, and the way in which the RDCs were incorporated as constraints in the protocol refinement, were consistent. We also tested the influence of different sets of RDCs and the best means of optimizing the calculation of D(a) and R. Resolution and accuracy of the ten best energy final structures were compared. The addition of a small set of RDC constraints significantly improves the final determined structures. We took advantage of the specificity of the RDC, i.e. it contains orientational information, and explored the global shape of the DNA duplexes; it was found that the duplexes do not have a large curvature. For the G-T base pair, we observed, in this particular sequence (tandem of G-T mismatches), a new pattern of base pairing, which involved the formation of a bifurcated hydrogen bond.

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Pre-organized structure of viral DNA at the binding-processing site of HIV-1 integrase

Cited by 23 publications

References 75 publications

Integration Requires a Specific Interaction of the Donor DNA Terminal 5′-Cytosine with Glutamine 148 of the HIV-1 Integrase Flexible Loop

Integration Requires a Specific Interaction of the Donor DNA Terminal 5′-Cytosine with Glutamine 148 of the HIV-1 Integrase Flexible Loop

Probing of HIV-1 Integrase/DNA Interactions Using Novel Analogs of Viral DNA

NMR assessment of the global shape of a non‐labelled DNA dodecamer containing a tandem of GT mismatches

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