User:Nathan Roy: Difference between revisions

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Once Gag is localized to discreet sites on the plasma membrane, multimerization of Gag takes place quite quickly , driven by the CA domain, and more specifically our focus here, the C-terminal domain of CA (CTD). There are four helices that contribute to the interaction of CA CTD with it's partner. A side-by-side interaction has been proposed (Figure 3), but many believe the forces involved in the side-by-side model are not great enough to account for the organization and structural stability of assembled Gag. Also, helix 1 of the CA CTD contains a very conserved region of residues within many retroviruses called the MHR (major homology region)<scene name='User:Nathan_Roy/Mhr_side-by-side/1'>MHR region</scene>. In the side-by-side model, the MHR is not responsible for the dimer organization, therefore a model of the CA CTD dimer in which the MHR is responsible for organization of the CA CTD dimers has been sought. By making a single deletion of the Ala 177 residue (which lies in the loop between helix 1 and helix 2), the CA CTD domain adopts a domain-swapped conformation,<applet load='2ont_dimer.pdb' size='300' frame='true' align='right' caption='FIGURE 4. Domain swapped CA CTD' /> in which the MHR of helix 1 is extended to contact helices 2,3, and 4 of the adjacent CA CTD domain (Figure 4)(<scene name='User:Nathan_Roy/Mhr_domain_swapped/1'>MHR region</scene>). ~~This domain swapping allows for a tighter binging~~ of the CA ~~CTD domains~~, ~~and a stronger, and more rigid~~ viral capsid.

Once Gag is localized to discreet sites on the plasma membrane, multimerization of Gag takes place quite quickly , driven by the CA domain, and more specifically our focus here, the C-terminal domain of CA (CTD). There are four helices that contribute to the interaction of CA CTD with it's partner. A side-by-side interaction has been proposed (Figure 3), but many believe the forces involved in the side-by-side model are not great enough to account for the organization and structural stability of assembled Gag. Also, helix 1 of the CA CTD contains a very conserved region of residues within many retroviruses called the MHR (major homology region)<scene name='User:Nathan_Roy/Mhr_side-by-side/1'>MHR region</scene>. In the side-by-side model, the MHR is not responsible for the dimer organization, therefore a model of the CA CTD dimer in which the MHR is responsible for organization of the CA CTD dimers has been sought. By making a single deletion of the Ala 177 residue (which lies in the loop between helix 1 and helix 2), the CA CTD domain adopts a domain-swapped conformation,<applet load='2ont_dimer.pdb' size='300' frame='true' align='right' caption='FIGURE 4. Domain swapped CA CTD' /> in which the MHR of helix 1 is extended to contact helices 2,3, and 4 of the adjacent CA CTD domain (Figure 4)(<scene name='User:Nathan_Roy/Mhr_domain_swapped/1'>MHR region</scene>). Keep in mind these are only models of Gag dimerization and subsequent multimerization, and that precise interactions that take place during Gag assembly are yet to be agreed upon.

Note the CA NTD was not discussed here. Through biochemical studies, the NTD domain has been implicated in viral capsid assembly, but reliable structural data giving biological insights has been elusive.

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As we have seen, Gag is responsible for correct targeting of viral assembly to discreet sites on the plasma membrane, and viral capsid structure assembly into organized virus particles. However, Gag is also responsible for packaging of the viral RNA into budding virions, and this function in executed by the NC domain. The 5' LTR of HIV-1 genomic RNA contains a recognition element called the psi element. All retroviruses contain some type of psi element in order to get specific packaging of viral genomic RNA within the budding particles, and in the case of HIV-1, the psi element is 120 bases, and contains 4 stem-loop structures. Although the psi element can be quite variable, the SL3 loop is highly conserved within HIV-1 strains. Figure 5 shows the NC domain of HIV-1 NL4-3 in complex with the SL3 loop of the viral psi element.

There are two zinc knuckle domains, with the zinc ion held by three Cys residues and a His(<scene name='User:Nathan_Roy/Zinc_knuckle/1'>Knuckle</scene>). Notice that guanine210 and guanine212 of the RNA interact with the F2 and F1 CCHC zinc knuckles respectively. In the F1 knuckle, G9 fits into a hydrophobic cleft formed by Val13, Phe16, Ile24, and Ala 25. G9 satisfies Watson-Crick hydrogen bonding by interacting with the NH groups from Phe16 and Ala25(<scene name='User:Nathan_Roy/F1_interaction/1'>Show residues</scene>), and also the CO group of Lys14

(<scene name='User:Nathan_Roy/F1_interaction/2'>Show</scene>). G7 interacts much the same with the F2 knuckle by hydrogen bonding with the NH groups of Trp37 and Met46, while also hydrogen bonding with the CO group of Gly35. The adenine211 nucleotide forms a hydrogen bond with a highly conserved Arg32 residue(<scene name='User:Nathan_Roy/F1_interaction/3'>Show</scene>). Also, residues 3 thru 10 form a 3.10 helix, which nestles into the RNA major groove(<scene name='User:Nathan_Roy/3-10_helix/1'>Show helix</scene>). These interactions allow viral RNA to be specifically packaged into virions.

(<scene name='User:Nathan_Roy/F1_interaction/2'>Show</scene>). G7 interacts much the same with the F2 knuckle by hydrogen bonding with the NH groups of Trp37 and Met46, while also hydrogen bonding with the CO group of Gly35. The adenine211 nucleotide forms a hydrogen bond with a highly conserved Arg32 residue(<scene name='User:Nathan_Roy/F1_interaction/3'>Show</scene>). Also, residues 3 thru 10 form a 3.10 helix, which nestles into the RNA major groove(<scene name='User:Nathan_Roy/3-10_helix/1'>Show helix</scene>). These interactions allow viral RNA to be specifically packaged into virions. Also of note, it has been shown that NC binding of viral RNA increases the ability of Gag to multimerize, thus providing another mechanism to couple functional virion assembly to Gag multimerization and vial budding.

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@@ Line 43: / Line 43: @@
 <applet load='1aum_dimer.pdb' size='300' frame='true' align='left' caption='FIGURE 3. Side-by-side structure of CA CTD' />
-Once Gag is localized to discreet sites on the plasma membrane, multimerization of Gag takes place quite quickly , driven by the CA domain, and more specifically our focus here, the C-terminal domain of CA (CTD). There are four helices that contribute to the interaction of CA CTD with it's partner. A side-by-side interaction has been proposed (Figure 3), but many believe the forces involved in the side-by-side model are not great enough to account for the organization and structural stability of assembled Gag. Also, helix 1 of the CA CTD contains a very conserved region of residues within many retroviruses called the MHR (major homology region)<scene name='User:Nathan_Roy/Mhr_side-by-side/1'>MHR region</scene>. In the side-by-side model, the MHR is not responsible for the dimer organization, therefore a model of the CA CTD dimer in which the MHR is responsible for organization of the CA CTD dimers has been sought.  By making a single deletion of the Ala 177 residue (which lies in the loop between helix 1 and helix 2), the CA CTD domain adopts a domain-swapped conformation,<applet load='2ont_dimer.pdb' size='300' frame='true' align='right' caption='FIGURE 4. Domain swapped CA CTD' /> in which the MHR of helix 1 is extended to contact helices 2,3, and 4 of the adjacent CA CTD domain (Figure 4)(<scene name='User:Nathan_Roy/Mhr_domain_swapped/1'>MHR region</scene>). This domain swapping allows for a tighter binging of the CA CTD domains, and a stronger, and more rigid viral capsid.
+Once Gag is localized to discreet sites on the plasma membrane, multimerization of Gag takes place quite quickly , driven by the CA domain, and more specifically our focus here, the C-terminal domain of CA (CTD). There are four helices that contribute to the interaction of CA CTD with it's partner. A side-by-side interaction has been proposed (Figure 3), but many believe the forces involved in the side-by-side model are not great enough to account for the organization and structural stability of assembled Gag. Also, helix 1 of the CA CTD contains a very conserved region of residues within many retroviruses called the MHR (major homology region)<scene name='User:Nathan_Roy/Mhr_side-by-side/1'>MHR region</scene>. In the side-by-side model, the MHR is not responsible for the dimer organization, therefore a model of the CA CTD dimer in which the MHR is responsible for organization of the CA CTD dimers has been sought.  By making a single deletion of the Ala 177 residue (which lies in the loop between helix 1 and helix 2), the CA CTD domain adopts a domain-swapped conformation,<applet load='2ont_dimer.pdb' size='300' frame='true' align='right' caption='FIGURE 4. Domain swapped CA CTD' /> in which the MHR of helix 1 is extended to contact helices 2,3, and 4 of the adjacent CA CTD domain (Figure 4)(<scene name='User:Nathan_Roy/Mhr_domain_swapped/1'>MHR region</scene>). Keep in mind these are only models of Gag dimerization and subsequent multimerization, and that precise interactions that take place during Gag assembly are yet to be agreed upon.
+Note the CA NTD was not discussed here. Through biochemical studies, the NTD domain has been implicated in viral capsid assembly, but reliable structural data giving biological insights has been elusive.
@@ Line 80: / Line 83: @@
 As we have seen, Gag is responsible for correct targeting of viral assembly to discreet sites on the plasma membrane, and viral capsid structure assembly into organized virus particles. However, Gag is also responsible for packaging of the viral RNA into budding virions, and this function in executed by the NC domain. The 5' LTR of HIV-1 genomic RNA contains a recognition element called the psi element. All retroviruses contain some type of psi element in order to get specific packaging of viral genomic RNA within the budding particles, and in the case of HIV-1, the psi element is 120 bases, and contains 4 stem-loop structures. Although the psi element can be quite variable, the SL3 loop is highly conserved within HIV-1 strains. Figure 5 shows the NC domain of HIV-1 NL4-3 in complex with the SL3 loop of the viral psi element.
 There are two zinc knuckle domains, with the zinc ion held by three Cys residues and a His(<scene name='User:Nathan_Roy/Zinc_knuckle/1'>Knuckle</scene>). Notice that guanine210 and guanine212 of the RNA interact with the F2 and F1 CCHC zinc knuckles respectively. In the F1 knuckle, G9 fits into a hydrophobic cleft formed by Val13, Phe16, Ile24, and Ala 25. G9 satisfies Watson-Crick hydrogen bonding by interacting with the NH groups from Phe16 and Ala25(<scene name='User:Nathan_Roy/F1_interaction/1'>Show residues</scene>), and also the CO group of Lys14
-(<scene name='User:Nathan_Roy/F1_interaction/2'>Show</scene>). G7 interacts much the same with the F2 knuckle by hydrogen bonding with the NH groups of Trp37 and Met46, while also hydrogen bonding with the CO group of Gly35. The adenine211 nucleotide forms a hydrogen bond with a highly conserved Arg32 residue(<scene name='User:Nathan_Roy/F1_interaction/3'>Show</scene>). Also, residues 3 thru 10 form a 3.10 helix, which nestles into the RNA major groove(<scene name='User:Nathan_Roy/3-10_helix/1'>Show helix</scene>). These interactions allow viral RNA to be specifically packaged into virions.
+(<scene name='User:Nathan_Roy/F1_interaction/2'>Show</scene>). G7 interacts much the same with the F2 knuckle by hydrogen bonding with the NH groups of Trp37 and Met46, while also hydrogen bonding with the CO group of Gly35. The adenine211 nucleotide forms a hydrogen bond with a highly conserved Arg32 residue(<scene name='User:Nathan_Roy/F1_interaction/3'>Show</scene>). Also, residues 3 thru 10 form a 3.10 helix, which nestles into the RNA major groove(<scene name='User:Nathan_Roy/3-10_helix/1'>Show helix</scene>). These interactions allow viral RNA to be specifically packaged into virions. Also of note, it has been shown that NC binding of viral RNA increases the ability of Gag to multimerize, thus providing another mechanism to couple functional virion assembly to Gag multimerization and vial budding.