Difference between revisions of "Part:BBa K404315"
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| + | <h2>Tymidine kinase</h2> | ||
| + | Thymidine kinase (TK) (EC 2.7.1.21) is known to be involved in the salvage pathway of nucleosides to nucleotides (Andrei et al. 2005). Due to its broader spectrum for different substrates, herpes simplex virus thymidine kinase (HSV-TK) is widely used in gene therapy approaches instead of endogenous thymidine kinases (Black et al. 1996). The transgenic introduced HSV-TK monophosphrylates nucleosides or nucleoside analogs such as ganciclovir (GCV) or acyclovir (AVC) followed by further phosphorylation through cellular kinases to nucleoside triphsphosphates. Incorporation of nucleotide analogs such as ganciclovir triphosphate or acyclovir triphosphates leads to DNA chain termination (Reardon 1989) and finally results in cell death. Genetic modifications of the active site represented by a tripeptide motif in thymidine kinase increases the substrate affinity of HSV-TK towards GCV and ACV (Black et al. 1996). Two promising mutant HSV-TKs have been found by large mutagenesis screenings modifying several amino acids and conducting sensitivity assays for ganciclovir and acyclovir (Black et al. 2001). | ||
| + | SR39 | ||
| + | In contrast to the TK30 mutant, the modified thymidine kinase SR39 obtained from a semi-random (SR) mutagenesis screening contains five modifications listed in Table 1 and provides further specificity for nucleoside analogs ganciclovir and acyclovir. <br> | ||
| + | <h2>Mouse guanylate kinase</h2> | ||
| + | Guanylate kinases (GMKs) are involved in the salvage pathway of mono-phosphorylated guanosine (GMP) nucleosides to GDP therefore being essential in nucleotide maturation (Stolworthy & Black 2001). By introducing transgenic thymidine kinases (TKs) into tumor cells, a bottleneck occurs by overexpression of mono-phosphorylated intermediates. To overcome the accumulation of these non-toxic molecules, Willmon, Krabbenhoft, & Black (2006) fused the herpes simplex virus thymidine kinase (HSV-TK) to the guanylate kinase from M. musculus and demonstrated enhanced tumor killing in vitro.<br> | ||
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Revision as of 00:42, 28 October 2010
Mouse guanylate kinase - tymidine kinase SR39 (Fusion protein mGMK_SR39)
Tymidine kinase
Thymidine kinase (TK) (EC 2.7.1.21) is known to be involved in the salvage pathway of nucleosides to nucleotides (Andrei et al. 2005). Due to its broader spectrum for different substrates, herpes simplex virus thymidine kinase (HSV-TK) is widely used in gene therapy approaches instead of endogenous thymidine kinases (Black et al. 1996). The transgenic introduced HSV-TK monophosphrylates nucleosides or nucleoside analogs such as ganciclovir (GCV) or acyclovir (AVC) followed by further phosphorylation through cellular kinases to nucleoside triphsphosphates. Incorporation of nucleotide analogs such as ganciclovir triphosphate or acyclovir triphosphates leads to DNA chain termination (Reardon 1989) and finally results in cell death. Genetic modifications of the active site represented by a tripeptide motif in thymidine kinase increases the substrate affinity of HSV-TK towards GCV and ACV (Black et al. 1996). Two promising mutant HSV-TKs have been found by large mutagenesis screenings modifying several amino acids and conducting sensitivity assays for ganciclovir and acyclovir (Black et al. 2001).
SR39
In contrast to the TK30 mutant, the modified thymidine kinase SR39 obtained from a semi-random (SR) mutagenesis screening contains five modifications listed in Table 1 and provides further specificity for nucleoside analogs ganciclovir and acyclovir.
Mouse guanylate kinase
Guanylate kinases (GMKs) are involved in the salvage pathway of mono-phosphorylated guanosine (GMP) nucleosides to GDP therefore being essential in nucleotide maturation (Stolworthy & Black 2001). By introducing transgenic thymidine kinases (TKs) into tumor cells, a bottleneck occurs by overexpression of mono-phosphorylated intermediates. To overcome the accumulation of these non-toxic molecules, Willmon, Krabbenhoft, & Black (2006) fused the herpes simplex virus thymidine kinase (HSV-TK) to the guanylate kinase from M. musculus and demonstrated enhanced tumor killing in vitro.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 653
Illegal NgoMIV site found at 1558 - 1000INCOMPATIBLE WITH RFC[1000]Illegal SapI.rc site found at 45