title: Nucleoside binding site of herpes simplex type 1 thymidine kinase analyzed by X-ray crystallography.
abstract: The crystal structures of the full-length Herpes simplex virus type 1 thymidine kinase in its unligated form and in a complex with an adenine analogue have been determined at 1.9 A resolution. The unligated enzyme contains four water molecules in the thymidine pocket and reveals a small induced fit on substrate binding. The structure of the ligated enzyme shows for the first time a bound adenine analogue after numerous complexes with thymine and guanine analogues have been reported. The adenine analogue constitutes a new lead compound for enzyme-prodrug gene therapy. In addition, the structure of mutant Q125N modifying the binding site of the natural substrate thymidine in complex with this substrate has been established at 2.5 A resolution. It reveals that neither the binding mode of thymidine nor the polypeptide backbone conformation is altered, except that the two major hydrogen bonds to thymidine are replaced by a single water-mediated hydrogen bond, which improves the relative acceptance of the prodrugs aciclovir and ganciclovir compared with the natural substrate. Accordingly, the mutant structure represents a first step toward improving the virus-directed enzyme-prodrug gene therapy by enzyme engineering.
authors.affiliation: Institut für Organische Chemie und Biochemie, Albert-Ludwigs-Universität, Freiburg im Breisgau, Germany.
authors.forename: J
authors.initials: J
authors.lastname: Vogt
authors.forename: R
authors.initials: R
authors.lastname: Perozzo
authors.forename: A
authors.initials: A
authors.lastname: Pautsch
authors.forename: A
authors.initials: A
authors.lastname: Prota
authors.forename: P
authors.initials: P
authors.lastname: Schelling
authors.forename: B
authors.initials: B
authors.lastname: Pilger
authors.forename: G
authors.initials: G
authors.lastname: Folkers
authors.forename: L
authors.initials: L
authors.lastname: Scapozza
authors.forename: G E
authors.initials: GE
authors.lastname: Schulz
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D000241:Adenosine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D000249:Adenosine Monophosphate / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D019943:Amino Acid Substitution; D000998:Antiviral Agents / Q000737:chemistry / Q000378:metabolism; D001665:Binding Sites; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000378:metabolism; D018259:Herpesvirus 1, Human / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D009154:Mutation; D009705:Nucleosides / Q000737:chemistry / Q000378:metabolism*; D063065:Organophosphonates; D011355:Prodrugs / Q000737:chemistry / Q000378:metabolism; D017434:Protein Structure, Tertiary; D013237:Stereoisomerism; D013379:Substrate Specificity; D013936:Thymidine / Q000737:chemistry / Q000378:metabolism; D013937:Thymidine Kinase / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism; D014764:Viral Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D014867:Water / Q000378:metabolism
chemical_list: C418964:9-(2-hydroxypropyl)adenine; D000998:Antiviral Agents; D004791:Enzyme Inhibitors; D009705:Nucleosides; D063065:Organophosphonates; D011355:Prodrugs; D014764:Viral Proteins; D014867:Water; D000249:Adenosine Monophosphate; C053001:adefovir; D013937:Thymidine Kinase; D000225:Adenine; D000241:Adenosine; D013936:Thymidine