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Introduction

The 1ki4 protein is the thymidine kinase from Herpes simplex virus type I complexed with 5-bromothienyldeoxyuridine. The Herpes simplex virus type I also called HSV1 cause highly contagious infections worldwide. In most cases, it is orofacial herpes but there is also a small proportion of genital infections.

The thymidine kinase is located in human cells and virus cells like in Herpes simplex virus. In the human cells, it exists two forms called TK1 and TK2.

This protein is a phosphotransferase enzyme that catalyses the reaction:

Deoxythymidine + ATP --> Deoxythymidine monophosphate + ADP

Function

The role of kinase is to catalyse protein phosphorylation. To do this, they transmit a phosphate group to the protein in the side chain of an amino acid chains. The phosphate group comes from the hydrolysis of ATP in ADP. This reaction can be in both directions.^[1]

The HSV1-TK is involved in the reactivation of Herpes simplex virus. Indeed this protein is involved in the salvage pathway of pyrimidine synthesis. As seen previously, this enzyme allows to transfert the γ phosphate group from ATP to the deoxythymidine (dT) to generate deoxythymidine monophosphate (dTMP).^[2]

Structural highlights

The HSV1-TK is a dimeric protein. The dimers have the same sequence because they come from the same gene, the gene TK.^[3] Each dimer has 331 amino acid residues^[4] and it has a molecular weight equal to 35,78 Da^[5]. Each chain is a polypeptide(L) which contains 50% α-helices and 11% β-sheets. The A chain has 18 α-helices with 167 residues and 9 β-sheets with 39 residues. Whereas the B chain has 19 α-helices with 168 residues and 9 β-sheets with 39 residues.^[6] The total molecular weight of the enzyme is equal to 72528 Da.^[7]

Each chain is bound to 2 chemical ligands. One of them is the sulfate ion SO4 which is linked to Gly 59, Gly 61, Lys 62, The 63 and Arg 222.^[8] The second one is the 5-Bromothienyleoxyuridine (BTD) which is linked to more amino acid^[9] as Trp 88, Gln 125, Tyr 172, Glu 225 and Tir 101^[10].

The active site is at the level of C-terminus of the first β-sheet of the polypeptide. It is surrounded by the lateral chain of 4 α-helices. It contains the thymidine which is in depth in the active site of the enzyme. It interacts with the enzyme thanks to Van der Waals interactions and hydrogen bond. In the periphery of the enzyme and so of the active site there is the binding site of ATP. The sulfate ion is nearby.^[11]

Disease

The HSV1-TK is located in Herpes simplex type I virus and causes orofacial infections.

Most of the time the orofacial herpes is asymptomatic and and carriers of the infection are unaware of it. Symptoms are vesicular lesions or painful open wounds (burning, itching…) inside the mouth.

HSV1 is transmitted mainly by contact with mucous membranes because of the presence of viral particles in wounds, saliva… It can also be transmitted to the genital sphere during oral sex, which causes genital herpes. In rare cases, the mother can transmit the virus to the new-born during the childbirth. In immunocompromised people, symptoms may be more severe and more frequent.

After the first infection, wounds can reappear, and the frequency depends on person. Antivirals are used to reduce the viral load of people infected but they do not cure the infection. However, a mutation in the HSV1-TK protein causes the resistance of the virus against antivirals in immunocompromised patients. ^[12]

Therapeutic application

The first suicide gene system developed for a therapeutic gene use, uses the thymidine kinase from Herpes simplex type I virus.

The aim was to develop a gene encoding an enzyme that performs an activity normally absent in the target cells of the therapy. This enzyme would then allow the synthesis of toxic compounds.

Aciclovir (ACV) and Ganciclovir (GVC) are 2 nucleosides analogues of guanosine. These are metabolized by HSV1-TK to a monophosphoryl compound which is then transformed into di and tri phosphate metabolites by cellular kinases. Then, ACV-TP and GVC-TP are incorporated into DNA during the elongation causing the death of infected cells by apoptosis. They also inhibit the viral replication. Death of cells is also triggered when they are treated with GVC when there has been an upstream transfection of the HSV1-TK gene outside a viral infection. ^[13]