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Computer-aided active-site-directed modeling of the herpes simplex virus 1 and human thymidine kinase
G Folkers1, S Trumpp-Kallmeyer, O Gutbrod
1Pharmaceutical Institute, University of Tübingen, Germany.
Journal of Computer-Aided Molecular Design
|October 1, 1991
Summary
Herpes Simplex Virus 1 thymidine kinase (HSV1-TK) and human TK have distinct active sites, explaining differing drug affinities. Mutations in HSV1-TK
Area of Science:
- Biochemistry
- Structural Biology
- Virology
Background:
- Thymidine kinase (TK) enzymes are crucial for antiviral drug efficacy, particularly for guanine derivatives like aciclovir (ACV).
- Herpes Simplex Virus 1 thymidine kinase (HSV1-TK) exhibits higher affinity for ACV than human TK, impacting antiviral treatment.
- Understanding the molecular basis of substrate binding differences is key to developing more effective antiviral therapies.
Purpose of the Study:
- To elucidate the structural differences in the active sites of HSV1-TK and human TK at a molecular level.
- To investigate the substrate binding interactions of thymidine and antiviral drugs with both HSV1-TK and human TK.
- To validate structural models through site-directed mutagenesis and enzymatic activity assays.
Main Methods:
- Development of three-dimensional (3-D) structural models for the active sites of HSV1-TK and human TK based on primary and secondary structure analysis.
- Validation of models using homologous enzymes with known 3-D structures.
- Construction and analysis of enzyme-substrate complexes (thymidine and dihydroxypropoxyguanine) using docking simulations and X-ray crystallography data (EF-Tu) as reference.
- Site-directed mutagenesis of HSV1-TK (Asp162 to Asn) to probe the thymidine recognition site.
Main Results:
- Both HSV1-TK and human TK models reveal a conserved core beta-sheet structure with nucleotide-binding motifs, similar to other kinases.
- Amino acid substitutions in homologous regions of HSV1-TK and human TK explain their differing substrate specificities.
- Site-directed mutagenesis of HSV1-TK at residue Asp162 abolished enzymatic activity, confirming its critical role in thymidine recognition.
Conclusions:
- The structural models provide insights into the molecular basis of substrate specificity differences between HSV1-TK and human TK.
- Specific amino acid variations within the active sites dictate the differential binding affinities for antiviral drugs.
- Targeting these structural differences could lead to the development of more selective and potent antiviral agents.