Team:Warwick/Safety

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Latest revision as of 09:35, 7 August 2014

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dont delete this. - waq RNA dependent RNA polymerase (RdRp) RdRp is an enzyme which catalyzes the replication of RNA from an RNA template, an essential protein for all viruses with an RNA genome. RdRp in Hepatitis C virus is also referred to as the NS5B protein, with a total molecular weight of 65 kDa. Heterelogous expression of NS5B has been achieved in insect and bacterial hosts, performing RNA-dependent RNA synthesis (Behrens et al., 1996; Lohmann et al., 1997). Hydrophobic C-terminal 21 amino acid residues cause insertion into the membrane, essential for HCV RNA replication (Moradpour et al., 2004). Amino acid substitution experiments of the 21 amino acid insertion sequence indicate functions beyond a membrane anchor role, with intracellular protein-protein interactions implicated (Moradpour et al., 2004).C-terminal tail preceding the C-terminal hydrophobic insertion sequence interacts with structural elements including the β-hairpin loop of NS5b (Leveque et al., 2003). The β-hairpin loop is believed to position the 3' terminus of the HCV viral RNA for correct initiation of viral RNA synthesis (Hong et al., 2001). RdRp initiates viral RNA synthesis with nucleotidyl transfer activity found within palm motifs A and C, with several amino acid residues implicated in NTP triphosphate contact (Bressanelli et al., 2002). NS5B activity has been demonstrated in vitro, with synthesis of full length HCV RNA (Lohmann et al., 1997; Ferrari et al., 1999). The 5’ and 3’ untranslated regions (UTRs) of the HCV genome contain ordered RNA structures, which are evolutionary conserved and contain crucial cis-acting elements for viral RNA replication. 150 nt in the 3’ termini of HCV RNA contains elements which are essential for RdRp binding and replication of viral RNA (Cheng et al., 1999; Yi and Lemon, 2003). http://www.ncbi.nlm.nih.gov/books/NBK1629/#ch10.r51 i

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+RNA dependent RNA polymerase (RdRp)
-Our modelling in this project has several aims :
+RdRp is an enzyme which catalyzes the replication of RNA from an RNA template, an essential protein for all viruses with an RNA genome. RdRp in Hepatitis C virus is also referred to as the NS5B protein, with a total molecular weight of 65 kDa. Heterelogous expression of NS5B has been achieved in insect and bacterial hosts, performing RNA-dependent RNA synthesis (Behrens et al., 1996; Lohmann et al., 1997). Hydrophobic C-terminal 21 amino acid residues cause insertion into the membrane, essential for HCV RNA replication (Moradpour et al., 2004). Amino acid substitution experiments of the 21 amino acid insertion sequence indicate functions beyond a membrane anchor role, with intracellular protein-protein interactions implicated (Moradpour et al., 2004).C-terminal tail preceding the C-terminal hydrophobic insertion sequence interacts with structural elements including the β-hairpin loop of NS5b (Leveque et al., 2003). The β-hairpin loop is believed to position the 3' terminus of the HCV viral RNA for correct initiation of viral RNA synthesis (Hong et al., 2001).
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+RdRp initiates viral RNA synthesis with nucleotidyl transfer activity found within palm motifs A and C, with several amino acid residues implicated in NTP triphosphate contact (Bressanelli et al., 2002). NS5B activity has been demonstrated in vitro, with synthesis of full length HCV RNA (Lohmann et al., 1997; Ferrari et al., 1999). The 5’ and 3’ untranslated regions (UTRs) of the HCV genome contain ordered RNA structures, which are evolutionary conserved and contain crucial cis-acting elements for viral RNA replication. 150 nt in the 3’ termini of HCV RNA contains elements which are essential for RdRp binding and replication of viral RNA (Cheng et al., 1999; Yi and Lemon, 2003).
- <li>To find the amount of DPP-IV reduction reached when the system reaches equilibrium</li>
- <li>To find a way to control the level of DPP-IV reduction</li>
- <li>To find the minimum number of RdRps, replicons, etc to be initially transfected into the cell, which are required to achieve a steady state for the system</li>
- <li>To find out how long does it take for the system to reach equilibrium</li>
- <li>To find out the level of reduction we need to treat diabetes</li>
- <li>To find out how stable the system is (i.e. will the system only work in very specific situations, or in lots of different systems?)
-</ul>
-<p><p>
-We are currently using Simbiology in Matlab and Copasi to model the system. We are currently adapting several different models, which come from research into HCV replicons, to our system. If our models can be made to fit our experiments well, we may extend our project to try and find a way to control the level of DPP-IV which is reduced. In addition modelling the system will allow it to be better optimised in the future, and optimum values for constants such as the strength of the ribosome binding sites, and the number of siRNAs produced by each degradation, so that the effect of our biobrick can be optimised.<p><p>
-Initially we determined that our system should reach some equilibrium after a certain amount of time. This is because firstly, HCV is a successful virus, so the replicons should not completely degrade away as time goes to infinity.  Secondly, since there are only a finite amount of resources within the cell, the number of replicons in the system cannot keep increasing forever. This means either the number of replicons must tend towards a certain constant (constant with respect to time), or the number of replicons should tend towards oscillations. <p><p>
+http://www.ncbi.nlm.nih.gov/books/NBK1629/#ch10.r51
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