The Rational of Catalytic Activity of Herpes Simplex Virus Thymidine Kinase

Abstract

Most antiherpes therapies exploit the large substrate acceptance of herpes simplex virus type 1 thymidine kinase (TK HSV1 ) relative to the human isoenzyme. The enzyme selectively phosphorylates nucleoside analogs that can either inhibit viral DNA polymerase or cause toxic effects when incorporated into viral DNA. To relate structural properties of TK HSV1 ligands to their chemical reactivity we have carried out ab initio quantum chemistry calculations within the density functional theory framework in combination with biochemical studies. Calculations have focused on a set of ligands carrying a representative set of the large spectrum of sugar-mimicking moieties and for which structural information of the TK HSV1 -ligand complex is available. The k cat values of these ligands have been measured under the same experimental conditions using an UV spectrophotometric assay. The calculations point to the crucial role of electric dipole moment of ligands and its interaction with the negatively charged residue Glu 225 . A striking correlation is found between the energetics associated with this interaction and the k cat values measured under homogeneous conditions. This finding uncovers a fundamental aspect of the mechanism governing substrate diversity and catalytic turnover and thus represents a significant step toward the rational design of novel and powerful prodrugs for antiviral and TK HSV1 -linked suicide gene therapies.

Biochemical and Structural Characterization of (South)-Methanocarbathymidine That Specifically Inhibits Growth of Herpes Simplex Virus Type 1 Thymidine Kinase-transduced Osteosarcoma Cells

“…These structural data as well as the phosphorylation pattern analysis corroborate the recently published data on phosphorylation in whole cells (35). Moreover the pseudosugar moiety of both (S)-MCT and (N)-MCT maintains a favorable dipole moment effect toward the residue Glu-225 (52). By contrast, the narrow substrate specificity of the cellular thymidine kinase speaks for a lower plasticity of hTK1 in adapting non-natural substrates.…”

“…According to a Ramachandran analysis the model features 93.5% residues in most favored and 6.5% in additionally allowed regions (47). Only residue Arg-163 is in a disallowed region in both subunits probably because it participates in catalysis (27,(52)(53)(54)(55). Due to a lack of defined density presumably caused by high mobility, some parts of the protein could not be modeled.…”

Crystal Structure of Varicella Zoster Virus Thymidine Kinase

“…1b and 3b). By analogy with HSV1-tk, there are two pockets named P1 and P2, which recognize the base and sugar (-like) moieties, respectively (35). The P1 region contains an additional subpocket that contacts 5-substituent on the pyrimidine ring.…”

“…There is no obvious structural explanation for the selectivity against the fluoroarabinosyl and fluororibosyl derivatives. A quantum chemical study of the ribose and ribose-like moieties of a series of substrates with HSV1-tk suggest that there is a correlation between the electrical dipole of the sugar-like group and the catalytic activity of the enzyme (35). Thus, it could be that similar effects of conformation and/or polarity are seen with VZV-tk.…”

The Pharmaceutical Biochemistry Group: Where Pharmaceutical Chemistry Meets Biology and Drug Delivery

“…More recently it emerged as a suitable suicide gene to be used as safety tools for stem cell transplantation (SCT) within cancer therapy, [29] gene replacement therapy [30] and as a powerful reporter gene for in situ monitoring of protein expression and localization of stem cells. [31] Our research focuses on characterizing the molecular mechanism governing the broad substrate specificity of HSV1 TK compared to human cytosolic TK [32][33][34][35][36][37][38] in order to address the problem of resistance towards antiviral compounds and the clinical limitations such as immunogenicity and treatment incompatibility discovered during the clinical application of the suicide gene approach based on HSV1 TK/ ganciclovir [39] using the interdisciplinary approach of the group. An effort has been made in order to design and develop new PET tracers for monitoring cancer and stem cells in vivo [40][41][42][43] in collaboration with Prof. Ametamey of the ETHZ.…”