1. Rational Design of High Affinity Interaction Between CC Chemokine Binding Protein vCCI and CCL17/TARC
- Author
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Guan, Wenyan, Stark, Lauren E, Zhang, Ning, Bains, Arjan, Martinez, Airam, Dupureur, Cynthia M, Colvin, Michael E, and LiWang, Patricia J
- Subjects
Biochemistry and Cell Biology ,Biological Sciences ,Infectious Diseases ,5.1 Pharmaceuticals ,Chemokine CCL17 ,Humans ,Viral Proteins ,Protein Binding ,Molecular Dynamics Simulation ,Amino Acid Sequence ,Chemokines ,CC ,Mutation ,Medicinal and Biomolecular Chemistry ,Medical Biochemistry and Metabolomics ,Biochemistry & Molecular Biology ,Biochemistry and cell biology ,Medical biochemistry and metabolomics ,Medicinal and biomolecular chemistry - Abstract
The poxvirus-derived protein vCCI (viral CC chemokine inhibitor) binds almost all members of the CC chemokine family with nanomolar affinity, inhibiting their pro-inflammatory actions. Understanding the affinity and specificity of vCCI could lead to new anti-inflammatory therapeutics. CCL17, also known as TARC, is unusual among CC chemokines by having only micromolar binding to vCCI. We have used sequence analysis and molecular simulations to determine the cause of this weak binding, which identified several locations in CCL17 where mutations seemed likely to improve binding to vCCI. Based on the aforementioned analysis, we expressed and tested multiple mutants of CCL17. We found two single point mutants V44K and Q45R that increased binding affinity to vCCI by 2-3-fold and, in combination, further improved affinity by 7-fold. The CCL17 triple mutant G17R/V44K/Q45R yielded a Kd of 0.25 ± 0.13 μM, a 68-fold improvement in affinity compared to the complex with wild-type CCL17. A quadruple mutant G17R/V44K/Q45R/R57W showed high affinity (0.59 ± 0.09 μM) compared to the wild type but lower affinity than the triple mutant. This work demonstrates that sequence comparisons and molecular simulations can predict chemokine mutations that increase the level of binding to vCCI, an important first step in developing engineered chemokine inhibitors useful for anti-inflammatory therapy.
- Published
- 2024