The Role of Herpes Simplex Virus-1 Thymidine Kinase Alanine 168 in Substrate Specificity
نویسندگان
چکیده
Herpes simplex virus type 1 (HSV) thymidine kinase (TK) has been widely used in suicide gene therapy for the treatment of cancer due to its broad substrate specificity and the inability of the endogenous human TK to phosphorylate guanosine analogs such as ganciclovir (GCV). The basis of suicide gene therapy is the introduction of a gene that encodes a prodrug-activating enzyme into tumor cells. After administration, the prodrug is selectively converted to a toxic drug by the suicide gene product thereby bringing about the eradication of the cancer cells. A major drawback to this therapy is the low activity the enzyme displays towards the prodrugs, requiring high prodrug doses that result in adverse side effects. Earlier studies revealed two HSV TK variants (SR39 and mutant 30) derived by random mutagenesis with enhanced activities towards GCV in vitro and in vivo. While these mutants contain multiple amino acid substitutions, molecular modeling suggests that substitutions at alanine 168 (A168) may be responsible for the observed increase in prodrug sensitivity. To evaluate this, site-directed mutagenesis was used to individually substitute A168 with phenylalanine or tyrosine to reflect the mutations found in SR39 and mutant 30, respectively. Additionally, kinetic parameters and the ability of these mutants to sensitize tumor cells to GCV in comparison to wild-type thymidine kinase were determined.
منابع مشابه
PCR detection of thymidine kinase gen of latent herpes simplex Virus type 1 in mice trigeminal ganglia
Herpes simplex virus type 1 establishes a latent infection in the peripheral nervous system following primary infection. During latent infection, virus genome exhibit limited transcription, with the HSV LATs consistently detected in latency infected ganaglia. Following ocular infection viral latency develops in the trigeminal ganglia. In this study PCR has been used for detection of HSV-1 nuc...
متن کاملRestriction Enzyme Analysis of Thymidine Kinase Gene in Four Iranian Isolates of Herpes Simplex Virus Type 1
متن کامل
Selective abolishment of pyrimidine nucleoside kinase activity of herpes simplex virus type 1 thymidine kinase by mutation of alanine-167 to tyrosine.
Herpes simplex virus type 1 (HSV-1) encodes a thymidine kinase (TK) that markedly differs from mammalian nucleoside kinases in terms of substrate specificity. It recognizes both pyrimidine 2'-deoxynucleosides and a variety of purine nucleoside analogs. Based on a computer modeling study and in an attempt to modify this specificity, an HSV-1 TK mutant enzyme containing an alanine-to-tyrosine mut...
متن کاملThe Effects and Pharmacokinetics of Acyclovir in Neonates
Acyclovir (9-[2-hydroxyethoxymethyl] guanine) is an acyclic nucleoside analogue of guanosine which is a potent and selective antiviral agent. Acycloviris converted to the monophosphate by thymidine kinase the virus-specific form of this enzyme and is subsequently converted to the triphosphate by the host cell kinase. Acyclovir triphosphate inhibits viral DNA-polymerase terminating the chain...
متن کاملIn vitro Interaction of HSV-1 ORF P with Both Thymidine Kinase (TK) and an Unidentified Cellular Protein
Herpes simplex virus type-1 (HSV-1) is a neurotropic pathogen of humans that establishes latent infection in the sensory ganglia innervating the site of primary infection. A number of genes control HSV-1 pathogenicity and latency. Open reading frame P (ORF P) is one of these genes that might have a role in latency and pathogenesis. A complication in the analysis of the role of ORF P in the HSV-...
متن کامل