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< | <StructureSection load='4NYF' size='350' side='right' caption='X-ray structure of the HIV-1 integrase catalytic core domain' scene='60/604477/Structure/1'>{{Sandbox_ESBS}}<!-- PLEASE ADD YOUR CONTENT BELOW HERE --> | ||
'''Dimer of HIV-1 integrase catalytic core domain''' is the active form of the '''3'-Processing''' reaction that occurs in patients cells suffering from '''AIDS'''. The catalytic core domain is just one of three parts composing the HIV-1 integrase. This enzyme performs mainly two specific reactions : the '''3'-processing''' and the '''integration of the viral DNA''' into the host genome. ''In vivo'' we can find this protein in several forms such as monomer, dimers and tetramers<ref name="Dells">PMID: 19091057</ref>. | '''Dimer of HIV-1 integrase catalytic core domain''' is the active form of the '''3'-Processing''' reaction that occurs in patients cells suffering from '''AIDS'''. The catalytic core domain is just one of three parts composing the HIV-1 integrase. This enzyme performs mainly two specific reactions : the '''3'-processing''' and the '''integration of the viral DNA''' into the host genome. ''In vivo'' we can find this protein in several forms such as monomer, dimers and tetramers<ref name="Dells">PMID: 19091057</ref>. | ||
==Biological role== | ==Biological role== | ||
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This protein is divided in three main domains: the N-terminal, the central and the C-terminal domain. | This protein is divided in three main domains: the N-terminal, the central and the C-terminal domain. | ||
The '''N-terminal domain''' presents a HHCC motif which is a pseudo zinc-finger complexing with zinc ions. The zinc ejection impedes the 3'-processing process and pertubs the integrase multimerisation<ref name="Carayon">PMID:20164093</ref>. Therefore the presence of this ion is necessary the virus life cycle. | The '''N-terminal domain''' (1-49) presents a HHCC motif which is a pseudo zinc-finger complexing with zinc ions. The zinc ejection impedes the 3'-processing process and pertubs the integrase multimerisation<ref name="Carayon">PMID:20164093</ref>. Therefore the presence of this ion is necessary for the virus life cycle. | ||
The '''central domain''' which corresponds to the catalytic domain contains the <scene name='60/604477/Catalytic_triad/1'>catalytic triad</scene> by association of two aspartates and one glutamate residues that coordinate bivalent ions, Cd++ in this structure but Mg++ or Mn++ ''in vivo'' <ref name="Liao">PMID:21426159</ref>. There is also an analogy of structure with the transposase core domain. This domain contains <scene name='60/604477/Structure/2'>residues</scene> between the 170-180 position involved in the packaging of the Uracil DNA glycosylase ('''UNG2''')<ref name="Zheng">PMID:23863879</ref> essential for the viral replication. | The '''central domain''' (50-213) which corresponds to the catalytic domain contains the <scene name='60/604477/Catalytic_triad/1'>catalytic triad</scene> by association of two aspartates and one glutamate residues that coordinate bivalent ions, Cd++ in this structure but Mg++ or Mn++ ''in vivo'' <ref name="Liao">PMID:21426159</ref>. There is also an analogy of structure with the transposase core domain and <scene name='60/604477/Residues_186-194/1'>residues 186-194</scene> help to interact with DNA by contact the major groove of viral and cellular DNA<ref name="Chen">PMID:10890912</ref>. This domain contains <scene name='60/604477/Structure/2'>residues</scene> between the 170-180 position involved in the packaging of the Uracil DNA glycosylase ('''UNG2''')<ref name="Zheng">PMID:23863879</ref> essential for the viral replication. | ||
The '''C-terminale domain''' allows the DNA binding in a non-specific manner and | The '''flexible elbow''' (195-220) is a 26-aa alpha-helix called <scene name='60/604477/Flexible_elbow/1'>alpha 6</scene> helix that links the C-ter domain to the catalytic core domain. It can be see like a flexible elbow because it offers conformation changes of the two previous domains during integration. Another property of this intermediary domain is the ability to contact the DNA phosphate backbone thanks to three main residues : K211, K215 and K219<ref name="Chen">PMID:10890912</ref>. | ||
The '''C-terminale domain''' (214-288) is a domain composed of five beta-sheets. It allows the DNA binding in a non-specific manner, HIV-1 IN dimer-dimer contacts, has a tethering role for DNA during the integration process and is also involved in the stability of the complex with DNA<ref name="Tsuruyama">PMID:23341952</ref>. | |||
The 3' processing reaction is mediated by integrase dimers formed at each viral DNA extremities. Further, these two dimers will form the integrase tetramer which is the active form for the concerted integration<ref name="Dells">PMID: 19091057</ref>. | |||
== DNA binding == | == DNA binding == | ||
The recognition of the viral DNA is an important step for the '''3' processing''' reaction. One of the key elements which performs the specific binding between the integrase and the cognate viral DNA is the <scene name='60/604477/Alpha4_helix/1'>alpha4 helix</scene> that is situated at the catalytic core surface | The recognition of the viral DNA, specifically the '''L'''ong-'''T'''erminal '''R'''epeat ('''LTR''') found at each end of proviral DNA, is an important step for the '''3' processing''' reaction. One of the key elements which performs the specific binding between the integrase and the cognate viral DNA is the <scene name='60/604477/Alpha4_helix/1'>alpha4 helix</scene> that is situated at the catalytic core surface<ref name="Hobaika">PMID:19808934</ref>. | ||
In order to have a tight binding, the integrase-DNA interaction requires optimal number of contacts such as hydrogen bonds, van der Waals forces and ionic interactions between DNA elements (bases, phosphates and sugars) and amino acid chains. Among the alpha4 helix residues, <scene name='60/604477/Lys_residues_alpha4_helix/2'>Lys156 and Lys159</scene> have a main role in the specific binding of integrase to DNA by forming bidentate hydrogen bonds which are higher specific than single hydrogen bonds<ref name="Hobaika">PMID:19808934</ref>. Moreover, by being positively charged, the lysine side chain provides ionic interactions as well. In addition, Lys-rich sequence implicated in this specific interaction include these two lysines. | |||
It has also be shown that <scene name='60/604477/Gln_alpha4_helix/2'>Gln148 residue</scene> contacts on adenine and cytosine of the 5' AC overhang in processed LTR<ref name="Hobaika">PMID:19808934</ref>. | |||
It also seems that there is a different sensitivity to the viral DNA sequence (LTR) depending on the presence of Mg++ or Mn++ suggesting that specific contacts between the integrase and the viral DNA extremity are more favored in a Mg++ containing environment<ref name="Carayon">PMID:20164093</ref>. | |||
== Function == | == Function == | ||
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- ''in vivo'' it catalyzes the '''transport the viral DNA into the nucleus''', it interacts with many proteins ('''VPR, VBP1, HIV-matrix,...''') and DNA to form the '''Pre-Integration Complex''' ('''PIC''') and allow the integration of the viral DNA into host genome and this mechanism can be divided in two reactions, the '''3'-Processing''' and the '''Strand Transfer'''<ref name="Dells">PMID: 19091057</ref> | - ''in vivo'' it catalyzes the '''transport the viral DNA into the nucleus''', it interacts with many proteins ('''VPR, VBP1, HIV-matrix,...''') and DNA to form the '''Pre-Integration Complex''' ('''PIC''') and allow the integration of the viral DNA into host genome and this mechanism can be divided in two reactions, the '''3'-Processing''' and the '''Strand Transfer'''<ref name="Dells">PMID: 19091057</ref> | ||
[[Image:3108751 idr-4-065f1.png | thumb | upright=3]] | |||
- ''in vitro'' researchers proved that two more reactions can be catalyzed by the integrase, the '''disintegration''' of the viral DNA and also a possible '''DNA polymerase activity'''<ref name="Liao">PMID:21426159</ref> to repair mismatches during integration. | - ''in vitro'' researchers proved that two more reactions can be catalyzed by the integrase, the '''disintegration''' of the viral DNA and also a possible '''DNA polymerase activity'''<ref name="Liao">PMID:21426159</ref> to repair mismatches during integration. | ||
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The 3'-processing is a reaction where the integrase and most precisely, the catalytic core by association with the N-ter domain catalyze the '''excision of two 3'-terminal nucleotides''' on both strands of the DNA due to a nucleophilic attack using water. Indeed it is a specific cleavage where two nucleotides on the 3' viral DNA extremities are excised, after that the two newly 3'-OH are used for a second reaction : the strand transfer. To make it possible, integrase needs Mg++ or Mn++ and also zinc ions. Moreover dimers of IN are formed to induce this reaction, then these two dimers are linked on the viral DNA extremities to form a tetramer of IN<ref name="Carayon">PMID:20164093</ref>. The 3' processing reaction occurs before the PIC penetration in the nucleus. | The 3'-processing is a reaction where the integrase and most precisely, the catalytic core by association with the N-ter domain catalyze the '''excision of two 3'-terminal nucleotides''' on both strands of the DNA due to a nucleophilic attack using water. Indeed it is a specific cleavage where two nucleotides on the 3' viral DNA extremities are excised, after that the two newly 3'-OH are used for a second reaction : the strand transfer. To make it possible, integrase needs Mg++ or Mn++ and also zinc ions. Moreover dimers of IN are formed to induce this reaction, then these two dimers are linked on the viral DNA extremities to form a tetramer of IN<ref name="Carayon">PMID:20164093</ref>. The 3' processing reaction occurs before the PIC penetration in the nucleus. | ||
===Strand Transfer=== | ===Strand Transfer=== | ||
The strand transfer is the really integration step, it provokes the integration of the viral DNA into the host genome by two sequential transesterifications. The integration site is a palindromic and symmetric sequence composed of five nucleotides. This sequence is cleaved to allow the fusion of the LTR extremities of the viral DNA. Afterwards two 3' nucleotides are excised and the joining strand of the five nucleotides is full-filed<ref name="Mbisa">PMID:21694910</ref>. The structure that permits this transfer is the intasome (viral DNA + integrase) by association with the '''Lens Epithelium-derived Growth factor (LEDGF)''' and the '''p75''' protein that has a cofactor activity for | The strand transfer is the really integration step, it provokes the integration of the viral DNA into the host genome by '''two sequential transesterifications'''. The integration site is a palindromic and symmetric sequence composed of five nucleotides. This sequence is cleaved to allow the fusion of the LTR extremities of the viral DNA. Afterwards two 3' nucleotides are excised and the joining strand of the five nucleotides is full-filed<ref name="Mbisa">PMID:21694910</ref>. The structure that permits this transfer is the intasome (viral DNA + integrase) by association with the '''Lens Epithelium-derived Growth factor (LEDGF)''' and the '''p75''' protein that has a cofactor activity for integrase and is a tethering site for chromatin binding. | ||
== Posttranslationnal Modifications == | == Posttranslationnal Modifications == | ||
Studies have prooved that 4 different kind of '''P'''ost '''T'''ranslationnal '''M'''odifications ('''PTMs''') affect the HIV-1 integrase : ubiquitination, SUMOylation, acetylation and phosphorylation. Furthermore, there are proteins that counteract or facilitate these PTMs implantation, for instance '''p300 and GCN5 can acetylate IN''' mostly on its C-ter domain on three different Lys residues (K264, K266 and K273). | |||
[[Image:1742-4690-7-18-7.jpg | thumb | upright=3]] | |||
In contrary '''Ku70 reduces the ubiquitination level''' of the N-ter domain (K6, K11, K27, K29, K33, K48 and K63). Another study also found that IN contains '''three ψ-K-x-D/E motifs''', which can be SUMOylated at three Lys residues, K46,<scene name='60/604477/Sumoylation/1'>K136</scene> and K244 <ref name="Terreni">PMID:20226045</ref>. | |||
==Inhibitors== | ==Inhibitors== | ||
Since few years, HIV-1 is an important therapeutic target. Actually there are two kind of inhibitors: the '''I'''ntegrase '''S'''trand '''T'''ransfer '''I'''nhibitors ('''INSTIs''' ) and the '''IN'''tegrase DNA-'''B'''inding '''I'''nhibitors '''(INBIs)'''. Over the past 5 years, INSTIs have been shown significant results as antiviral compounds with in 2007, the licensing of the first integrase inhibitor called '''raltegravir''', which target the integrase active site and thus, inhibit DNA strand transfer. However, resistance to this compound emerges which in turn confers the same effect to the second licensed INSTI, '''elvitegravir'''<ref name="Engelman">PMID: 23647983</ref>. | Since few years, HIV-1 is an important therapeutic target. Actually there are two kind of inhibitors: the '''I'''ntegrase '''S'''trand '''T'''ransfer '''I'''nhibitors ('''INSTIs''' ) and the '''IN'''tegrase DNA-'''B'''inding '''I'''nhibitors '''(INBIs)'''. Over the past 5 years, INSTIs have been shown significant results as antiviral compounds with in 2007, the licensing of the first integrase inhibitor called '''raltegravir''', which target the integrase active site and thus, inhibit DNA strand transfer. However, resistance to this compound emerges which in turn confers the same effect to the second licensed INSTI, '''elvitegravir'''<ref name="Engelman">PMID: 23647983</ref>. | ||
Recent advances allow the discovery of a new inhibitor class which acts in a dissimilar way from that of INSTIs. These compounds exert their inhibitory effect by binding to a highy conserved allosteric pocket on the catalytic core domain dimer ('''CCD-CCD dimer''') mediated through the alpha helice 1 and 5<ref name="Engelman">PMID:23647983</ref>. | Recent advances allow the discovery of a new inhibitor class which acts in a dissimilar way from that of INSTIs. These compounds exert their inhibitory effect by binding to a highy conserved allosteric pocket on the catalytic core domain dimer ('''CCD-CCD dimer''') mediated through the alpha helice 1 and 5<ref name="Engelman">PMID:23647983</ref>. | ||
In a normal HIV-1 life cycle, the cellular co-factor LEDGF/p75 engages the enzyme at the CCD-CCD dimer via its '''I'''ntegrase '''B'''inding '''D'''omain ('''IBD'''). By binding this allosteric pocket, LEDGF/p75 stimulates both concerted integration and strand transfer. It has also be shown that LEDGF-integrase complex exhibits a better solubility profiles compared to the free integrase. This co-factor increases the tetrameric stability of integrase in order to tether the | In a normal HIV-1 life cycle, the cellular co-factor '''LEDGF/p75''' engages the enzyme at the CCD-CCD dimer via its '''I'''ntegrase '''B'''inding '''D'''omain ('''IBD'''). By binding this allosteric pocket, LEDGF/p75 stimulates both concerted integration and strand transfer. It has also be shown that LEDGF-integrase complex exhibits a better solubility profiles compared to the free integrase. This co-factor increases the tetrameric stability of integrase in order to tether the viral DNA to active genes<ref name="Dells">PMID: 19091057</ref>. | ||
LEDGF/p75 contacts on one integrase subunit via hydrogen bonds between the cofactor residue '''Asp366''' and the primary amino groups of integrase residues <scene name='60/604477/ | LEDGF/p75 contacts on one integrase subunit via hydrogen bonds between the cofactor residue '''Asp366''' and the primary amino groups of integrase residues <scene name='60/604477/Glu170_his171/1'>Glu170 and His171</scene>. LEDGF/p75 residue '''Ile365''' contacts on the other integrase subunit by nesting into a hydrophobic pocket which consists of residues <scene name='60/604477/Leu_ala_trp_residues2/1'>Leu102, Ala128 and Trp132</scene><ref name="Engelman">PMID:23647983</ref>. | ||
This new class of inhibitor, by engaging this allosteric pocket, prevents the binding of LEDGF/p75 and inhibits the integrase activity by enhancing the formation of integrase multimers, which impedes the intasome assembly<ref name="Engelman">PMID:23647983</ref>. | This new class of inhibitor, by engaging this allosteric pocket, prevents the binding of LEDGF/p75 and inhibits the integrase activity by enhancing the formation of integrase multimers, which impedes the intasome assembly<ref name="Engelman">PMID:23647983</ref>. | ||