Herpesviral thymidine kinases: laxity and resistance by design.

Evans, James S.; Lock, Kevin P.; Levine, Barry A.; Champness, J.N.; Sanderson, Mark; Summers, William C.; McLeish, P.; Buchan, A.

doi:10.1099/0022-1317-79-9-2083

Cited by 34 publications

(24 citation statements)

References 28 publications

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“…Glu-83, Trp-88, Tyr-101, Gln-125, Arg-163, Tyr-172, and Glu-225 (HSV1-TK numbering), are conserved in DmdNK. These amino acids were also reported to be conserved in human TK2 (29).…”

Section: Sequencing Of An Estmentioning

confidence: 81%

Functional Expression of a Multisubstrate Deoxyribonucleoside Kinase from Drosophila melanogaster and Its C-terminal Deletion Mutants

Munch‐Petersen¹,

Knecht²,

Lenz³

et al. 2000

Journal of Biological Chemistry

116

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The occurrence of a deoxyribonucleoside kinase in Drosophila melanogaster (Dm-dNK) with remarkably broad substrate specificity has recently been indicated (Munch-Petersen, B., Piskur, J., and Søndergaard, L. (1998) J. Biol. Chem. 273, 3926 -3931). To prove that the capacity to phosphorylate all four deoxyribonucleosides is in fact associated to one polypeptide chain, partially sequenced cDNA clones, originating from the Berkeley Drosophila genome sequencing project, were searched for homology with human deoxyribonucleoside kinases. The total sequence of one cDNA clone and the corresponding genomic DNA was determined and expressed in Escherichia coli as a glutathione S-transferase fusion protein. The purified and thrombin cleaved recombinant protein phosphorylated the four deoxyribonucleosides with high turnover and K m values similar to those of the native Dm-dNK, as well as the four ribonucleosides and many therapeutical nucleoside analogs. DmdNK has apparently the same origin as the mammalian kinases, thymidine kinase 2, deoxycytidine kinase, deoxyguanosine kinase, and the herpesviral thymidine kinases, but it has a unique C terminus that seems to be important for catalytic activity and specificity. The Cterminal 20 amino acids were dispensable for phosphorylation of deoxyribonucleosides but necessary for full activity with purine ribonucleosides. Removal of the C-terminal 20 amino acids increased the specific activity 2-fold, but 99% of the activity was lost after removal of the C-terminal 30 amino acids.

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“…Glu-83, Trp-88, Tyr-101, Gln-125, Arg-163, Tyr-172, and Glu-225 (HSV1-TK numbering), are conserved in DmdNK. These amino acids were also reported to be conserved in human TK2 (29).…”

Section: Sequencing Of An Estmentioning

confidence: 81%

Functional Expression of a Multisubstrate Deoxyribonucleoside Kinase from Drosophila melanogaster and Its C-terminal Deletion Mutants

Munch‐Petersen¹,

Knecht²,

Lenz³

et al. 2000

Journal of Biological Chemistry

116

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“…Clusters of positive charges from the LID domain and the Mg 2ϩ make the phosphorus atom amenable for a nucleophilic attack of the 5Ј-OH of deoxyribose which is polarized by the two glutamines localized at positions 225 and 83 (239). The substrate binding site is composed of amino acid side chains derived primarily from a set of roughly parallel helices (85). These helices are localized at the N-terminal half of the polypeptide following the glycine-rich loop involved in the binding of the phosphate moiety of ATP.…”

Section: Mechanism Of Resistance To Nucleoside Analoguesmentioning

confidence: 99%

“…The most important sites involved in the enzyme activity are the ATP-binding site (aa 51 to 63), the nucleoside-binding site (aa 168 to 176), and the cysteine at codon 336 which maintains the three-dimensional structure of the active site (85 The structure of the viral TK consists of an ␣␤ structure made up of 15 ␣-helices and 7 ␤-sheets (36). Five-stranded parallel ␤-sheets form part of the core of the protein, which contains the active site.…”

Section: Mechanism Of Resistance To Nucleoside Analoguesmentioning

confidence: 99%

Resistance of Herpes Simplex Viruses to Nucleoside Analogues: Mechanisms, Prevalence, and Management

Piret

Boivin

2011

Antimicrob Agents Chemother

461

406

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“…Using a method previously described (4), it was confirmed that the Q125H mutation was not a part of natural polymorphism. Q125 of HSV-1 TK has been shown to be located above the nucleotide binding pocket in the three-dimensional (3D) structure of the vTK protein (12). Several studies have shown that substitution of Q125 to other amino acids changes vTK activity; Q125E and Q125L are associated with resistance to ACV, and Q125N leads to hypersensitivity to ACV (13,14).…”

mentioning

confidence: 99%

Neonatal Herpes Encephalitis Caused by a Virologically Confirmed Acyclovir-Resistant Herpes Simplex Virus 1 Strain

et al. 2013

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A neonate with herpes simplex virus 1 encephalitis was treated with intravenous acyclovir. During the course of therapy, the infection became intractable to the treatment and a mutation in the viral thymidine kinase gene (nucleotide G375T, amino acid Q125H) developed. This mutation was demonstrated in vitro to confer acyclovir resistance. CASE REPORTA 13-day-old boy was admitted to National Defense Medical College Hospital due to lethargy and failure to thrive. He was born at 39 weeks 5 days of gestation and 2,558 g birth weight to a healthy 35-year-old mother (gravida 2, para 2). Group B streptococcus (GBS) was detected from the mother's vagina in the third trimester, but the baby's bacterial culture tests performed at birth, including throat, skin, and blood analyses, were negative for GBS. The mother did not have a history of genital herpes. Her herpes simplex virus 1 (HSV-1) and HSV-2 serostatus was not examined, and her history of acyclovir (ACV) use was not clear. Furthermore, the genital swab culture examination for HSV was not performed. On admission, physical examination of the boy revealed skin blisters on the forehead and upper lip. A swab from the blister showed positive and negative reactions for the specific antigens of HSV-1 and HSV-2, respectively, in a direct immunofluorescent antibody assay (Denka Seiken Co. Ltd., Tokyo, Japan) performed by a qualified clinical laboratory (SRL Inc., Tokyo, Japan). A serum sample collected on admission showed positive and negative reactions in the enzyme-linked immunosorbent assay for detection of anti-HSV IgM and IgG antibody (SRL Inc.), respectively. A lumbar puncture revealed pleocytosis (547 cells/l) and an elevated protein level (168 mg/dl) in the cerebrospinal fluid (CSF). The CSF was also positive for HSV-1 DNA, which was determined by a previously reported method (1) in PCR testing (SRL Inc.). The boy was diagnosed as having neonatal herpes encephalitis (NHE), and intravenous highdose ACV (60 mg/kg/day) treatment was initiated. His general status improved with resolution of the skin lesions within a few days of the beginning of treatment. However, the viral load in the CSF determined by TaqMan-based quantitative real-time PCR (SRL Inc.), which dropped temporarily, increased again after 4 weeks from the initiation of ACV treatment (Fig. 1A) without obvious deterioration in clinical symptoms. Because the standard dose of ACV was given and drugs which have antagonistic effects for ACV were not used, we assumed that an ACV-resistant HSV-1 strain had developed. The ACV concentration in the CSF was not measured. Foscarnet, an antiviral drug recommended for treatment of ACV-resistant HSV infections (2), was not immediately available. Therefore, vidarabine (15 mg/kg/day) was added to the therapeutic regimen from the fifth week of the treatment course. Subsequently, HSV-DNA in the CSF decreased to a level that was finally undetectable; hence, the antiviral drug treatment was terminated. Because virus isolation from the CSF of the patient was unsuccessful, as is c...

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Herpesviral thymidine kinases: laxity and resistance by design.

Cited by 34 publications

References 28 publications

Functional Expression of a Multisubstrate Deoxyribonucleoside Kinase from Drosophila melanogaster and Its C-terminal Deletion Mutants

Functional Expression of a Multisubstrate Deoxyribonucleoside Kinase from Drosophila melanogaster and Its C-terminal Deletion Mutants

Resistance of Herpes Simplex Viruses to Nucleoside Analogues: Mechanisms, Prevalence, and Management

Neonatal Herpes Encephalitis Caused by a Virologically Confirmed Acyclovir-Resistant Herpes Simplex Virus 1 Strain

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