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 mutation at amino acid position 167 was constructed. Compared with wild-type HSV-1 TK, the purified mutant HSV-1 TK(A167Y) enzyme was heavily compromised in phosphorylating pyrimidine nucleosides such as (E)-5-(2-bromovinyl)-2'-deoxyuridine and the natural substrate dThd, whereas its ability to phosphorylate the purine nucleoside analogs ganciclovir (GCV) and lobucavir was only reduced approximately 2-fold. Moreover, a markedly decreased competition of natural pyrimidine nucleosides (i.e., thymidine) with purine nucleoside analogs for phosphorylation by HSV-1 TK(A167Y) was observed. Human osteosarcoma cells transduced with the wild-type HSV-1 TK gene were extremely sensitive to the cytostatic effects of antiherpetic pyrimidine [i.e., (E)-5-(2-bromovinyl)-2'-deoxyuridine] and purine (i.e., GCV) nucleoside analogs. Transduction with the HSV-1 TK(A167Y) gene sensitized the osteosarcoma cells to a variety of purine nucleoside analogs, whereas there was no measurable cytostatic activity of pyrimidine nucleoside analogs. The unique properties of the A167Y mutant HSV-1 TK may give this enzyme a therapeutic advantage in an in vivo setting due to the markedly reduced dThd competition with GCV for phosphorylation by the HSV-1 TK.

Degrève B ，Esnouf R ，De Clercq E ，Balzarini J ... -  《MOLECULAR PHARMACOLOGY》

The A167Y mutation converts the herpes simplex virus type 1 thymidine kinase into a guanosine analogue kinase.

Balzarini J ，Liekens S ，Esnouf R ，De Clercq E ... -  《-》

Mutation of Gln125 to Asn selectively abolishes the thymidylate kinase activity of herpes simplex virus type 1 thymidine kinase.

Degrève B ，Esnouf R ，De Clercq E ，Balzarini J ... -  《-》

Engineering of a single conserved amino acid residue of herpes simplex virus type 1 thymidine kinase allows a predominant shift from pyrimidine to purine nucleoside phosphorylation.

Balzarini J ，Liekens S ，Solaroli N ，El Omari K ，Stammers DK ，Karlsson A ... -  《-》

A new acycloguanosine-specific supermutant of herpes simplex virus type 1 thymidine kinase suitable for PET imaging and suicide gene therapy for potential use in patients treated with pyrimidine-based cytotoxic drugs.

Likar Y ，Dobrenkov K ，Olszewska M ，Vider E ，Shenker L ，Cai S ，Pillarsetty N ，Hricak H ，Ponomarev V ... -  《-》