Difference between revisions of "Part:BBa K404153"

Revision as of 14:12, 27 October 2010

[AAV2]-NLS

General informations

The AAV capsid consists of 60 capsid protein subunits composed of the three cap proteins VP1, VP2, and VP3, which are encoded in an overlapping reading frame. Arranged in a stoichiometric ratio of 1:1:10, they form an icosahedral symmetry. The mRNA encoding for the cap proteins is transcribed from p40 and alternative spliced to minor and major products. Alternative splicing and translation initiation of VP2 at a nonconventional ACG initiation codon promote the expression of the VP proteins. VP1, VP2 and VP3 share a common C terminus and stop codon, but begin with a different start codon. The N termini of VP1 and VP2 play important roles in infection and contain motifs that are highly homologous to a phospholipase A2 (PLA2) domain and nuclear localization signals (NLSs). These elements are conserved in almost all parvoviruses. (Johnson et al., 2010a).

Whereas VP1 is translated from the minor spliced mRNA, while VP2 and VP3 are translated from the major spliced mRNA. The minor spliced product is approximately 10-fold less abundant than the major spliced mRNA. Thus, there is much less VP1 than VP2 and VP3 resulting in a capsid stoichiometric ratio of 1:1:10. The N terminus of VP1 has an extension of 65 amino acids including an additional extension of 138 N-terminal amino acids forming the unique portion of VP1. It contains a motif of about 70 amino acids that is highly homologous to phospholipase A2 (PLA2) domain. Furthermore, there are nuclear localization sequences (BR)(+), which are supposed to be necessary for endosomal escape and nuclear entry. (Bleker, Pawlita, & Kleinschmidt, 2006), (DiPrimio, Asokan, Govindasamy, Agbandje-McKenna, & Samulski, 2008), (Johnson et al., 2010a).

The translation of VP2 from the major spliced mRNA is less efficient compared to the translation of VP3 because it initiates at Thr codon (ACG). VP2 and VP1 have an extension at the N terminus that remains internal when exposing the capsid to experimental conditions like low pH or heat. The exact role of VP2 remains unknown, although the protein is thought to be nonessential for viral assembly and infectivity. (DiPrimio, Asokan, Govindasamy, Agbandje-McKenna, & Samulski, 2008), (Johnson et al., 2010b)

NLS

NLS are located in basic regions on the N terminus of VP2 (35 aa) and VP1 (172 aa) and mediate genome delivery into the nucleus and transduction [Hoque et al.,1999; Grieger et al., 2006]. Nuclear localisation sequence is hydrophilic and contains ß-turn and coil regions [Kalderon, et al, 1984]. It was also described in CPV and MVM viruses. Compared to CPV, MVM virus contains several NLS within the capsid, which are activated at different infection stages [Lombardo et al., 2000; Lombardo et al., 2002]

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
COMPATIBLE WITH RFC[1000]

References

Bleker, S., Pawlita, M., & Kleinschmidt, J., 2006. IImpact of capsid conformation and Rep-capsid interactions on adeno-associated virus type 2 genome packaging. Journal of virology, 80(2), 810-820. doi: 10.1128/JVI.80.2.810.
DiPrimio, N., Asokan, A., Govindasamy, L., Agbandje-McKenna, M., & Samulski, R. J. , 2008. Surface loop dynamics in adeno-associated virus capsid assembly. Journal of virology, 82(11), 5178-89. doi: 10.1128/JVI.02721-07.
Hoque, 1999. Nuclear transport of the major capsid protein is essential for adeno-associated virus capsid formation. Journal of Virology, 73(9), pp.7912-7915.
Kalderon, D, 1984. Sequence requirements for nuclear location of simian virus 40 large-T antigen. Nature, 311(5981), pp.33-38
Johnson, J.S., 2010. Mutagenesis of adeno-associated virus type 2 capsid protein VP1 uncovers new roles for basic amino acids in trafficking and cell-specific transduction. Journal of virology, 84(17), 8888-902. doi: 10.1128/JVI.00687-10.
Lombardo, E, 2000. A beta-stranded motif drives capsid protein oligomers of the parvovirus minute virus of mice into the nucleus for viral assembly. Journal of Virology, 74(8), pp.3804-3814
Lombardo, E , 2002. Complementary roles of multiple nuclear targeting signals in the capsid proteins of the parvovirus minute virus of mice during assembly and onset of infection. Journal of Virology, 76(14), pp.7049-7059

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 __NOTOC__
 <partinfo>BBa_K404153 short</partinfo>
+<h3>General informations</h3>
+<html>
+<center>
+<img src=https://static.igem.org/mediawiki/2010/e/e8/Freiburg10_VP1-3_overview.jpg width=400>
+</center>
+</html><br>
+The AAV capsid consists of 60 capsid protein subunits composed of the three cap proteins VP1, VP2, and VP3, which are encoded in an overlapping reading frame. Arranged in a stoichiometric ratio of 1:1:10, they form an icosahedral symmetry. The mRNA encoding for the cap proteins is transcribed from p40 and alternative spliced to minor and major products. Alternative splicing and translation initiation of VP2 at a nonconventional ACG initiation codon promote the expression of the VP proteins. VP1, VP2 and VP3 share a common C terminus and stop codon, but begin with a different start codon. The N termini of VP1 and VP2 play important roles in infection and contain motifs that are highly homologous to a phospholipase A2 (PLA2) domain and nuclear localization signals (NLSs). These elements are conserved in almost all parvoviruses. (Johnson et al., 2010a). <br/>
+<br/>
+Whereas VP1 is translated from the minor spliced mRNA, while VP2 and VP3 are translated from the major spliced mRNA. The minor spliced product is approximately 10-fold less abundant than the major spliced mRNA. Thus, there is much less VP1 than VP2 and VP3 resulting in a capsid  stoichiometric ratio of 1:1:10. The N terminus of VP1 has an extension of 65 amino acids including an additional extension of 138 N-terminal amino acids forming the unique portion of VP1. It contains a motif of about 70 amino acids that is highly homologous to  phospholipase A2 (PLA2) domain.  Furthermore, there are nuclear localization sequences (BR)(+), which are supposed to be necessary for endosomal escape and nuclear entry. (Bleker, Pawlita, & Kleinschmidt, 2006), (DiPrimio, Asokan, Govindasamy, Agbandje-McKenna, & Samulski, 2008), (Johnson et al., 2010a). <br />
+<br/>
+The translation of VP2 from the major spliced mRNA is less efficient compared to the translation of VP3 because it initiates at  Thr codon (ACG). VP2 and VP1 have an extension at the N terminus that remains internal when exposing the capsid to experimental conditions like low pH or heat. The exact role of VP2 remains unknown, although the protein is thought to be nonessential for viral assembly and infectivity. (DiPrimio, Asokan, Govindasamy, Agbandje-McKenna, & Samulski, 2008), (Johnson et al., 2010b)
+<h3>NLS</h3>
 NLS are located in basic regions on the N terminus of VP2 (35 aa) and VP1 (172 aa) and mediate genome delivery into the nucleus and transduction [Hoque et al.,1999; Grieger et al., 2006]. Nuclear localisation sequence is hydrophilic and contains ß-turn and coil regions [Kalderon, et al, 1984]. It was also described in CPV and MVM viruses. Compared to CPV, MVM virus contains several NLS within the capsid, which are activated at different infection stages [Lombardo et al., 2000; Lombardo et al., 2002]
@@ Line 17: / Line 29: @@
 <!-- -->
 <h3>References</h3>
+<b>Bleker, S., Pawlita, M., & Kleinschmidt, J.</b>, 2006. IImpact of capsid conformation and Rep-capsid interactions on adeno-associated virus type 2 genome packaging. Journal of virology, 80(2), 810-820. doi: 10.1128/JVI.80.2.810. <br />
+<b>DiPrimio, N., Asokan, A., Govindasamy, L., Agbandje-McKenna, M., & Samulski, R. J. </b>, 2008. Surface loop dynamics in adeno-associated virus capsid assembly. Journal of virology, 82(11), 5178-89. doi: 10.1128/JVI.02721-07. <br />
 <b>Hoque</b>, 1999. Nuclear transport of the major capsid protein is essential for adeno-associated virus capsid formation. Journal of Virology, 73(9), pp.7912-7915. <br />
 <b>Kalderon, D</b>, 1984. Sequence requirements for nuclear location of simian virus 40 large-T antigen. Nature, 311(5981), pp.33-38 <br />
+<b>Johnson, J.S.</b>, 2010. Mutagenesis of adeno-associated virus type 2 capsid protein VP1 uncovers new roles for basic amino acids in trafficking and cell-specific transduction. Journal of virology, 84(17), 8888-902. doi: 10.1128/JVI.00687-10.<br />
 <b>Lombardo, E</b>, 2000. A beta-stranded motif drives capsid protein oligomers of the parvovirus minute virus of mice into the nucleus for viral assembly. Journal of Virology, 74(8), pp.3804-3814 <br />
 <b>Lombardo, E </b>, 2002. Complementary roles of multiple nuclear targeting signals in the capsid proteins of the parvovirus minute virus of mice during assembly and onset of infection. Journal of Virology, 76(14), pp.7049-7059