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Inhibition of a viral enzyme by a small-molecule dimer disruptor

Nature Chemical Biology volume 5, pages 640–646 (2009)Cite this article

Abstract

We identified small-molecule dimer disruptors that inhibit an essential dimeric protease of human Kaposi's sarcoma–associated herpesvirus (KSHV) by screening an α-helical mimetic library. Next, we synthesized a second generation of low-micromolar inhibitors with improved potency and solubility. Complementary methods including size exclusion chromatography and 1H-13C HSQC titration using selectively labeled 13C-Met samples revealed that monomeric protease is enriched in the presence of inhibitor. 1H-15N HSQC titration studies mapped the inhibitor binding site to the dimer interface, and mutagenesis studies targeting this region were consistent with a mechanism where inhibitor binding prevents dimerization through the conformational selection of a dynamic intermediate. These results validate the interface of herpesvirus proteases and other similar oligomeric interactions as suitable targets for the development of small-molecule inhibitors.

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Figure 1: KSHV Pr dimer interface and helical mimetic inhibitors of KSHV Pr activity.
Figure 2: DD2 disrupts the KSHV Pr dimer.
Figure 3: Kinetic studies with DD2 show evidence of mixed-type inhibition.
Figure 4: 1H-15N HSQC titration data map the DD2 binding site to the dimer interface.
Figure 5: Monomer trap model of inhibition by compound DD2.

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Acknowledgements

The authors thank W. Gibson (The Johns Hopkins University School of Medicine) for providing us with the CMV protease expression plasmid and for valuable feedback on the manuscript. This work was funded by US National Institutes of Health grants T32 GMO7810 and AIO67423 (C.S.C.) and by the American Lebanese and Syrian Associated Charities and the St. Jude Children's Research Hospital (R.K.G.). We also thank the University of California, San Francisco–Gladstone Institute for Virology and Immunology/Center for AIDS Research for the Clinical Science Pilot Award (P30-AI027763 to G.M.L.).

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Authors and Affiliations

  1. Graduate Group in Biochemistry and Molecular Biology, University of California, San Francisco, California, USA

    Tina Shahian

  2. Department of Pharmaceutical Chemistry, University of California, San Francisco, California, USA

    Gregory M Lee, Ana Lazic & Charles S Craik

  3. Department of Chemical Biology and Therapeutics, St. Jude Children's Research Hospital, Memphis, Tennessee, USA

    Leggy A Arnold, Priya Velusamy, Christina M Roels & R Kiplin Guy

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Contributions

T.S., G.M.L., A.L., C.S.C. and R.K.G. designed research; T.S., G.M.L. and A.L. carried out research; L.A.A., P.V. and C.M.R. performed chemical synthesis; T.S., G.M.L., A.L., C.S.C. and R.K.G. analyzed and interpreted data; and T.S., C.S.C. and R.K.G. prepared the manuscript.

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Correspondence to Charles S Craik.

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Supplementary Figures 1–4, Supplementary Scheme 1, Supplementary Tables 1 and 2, and Supplementary Methods (PDF 481 kb)

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Shahian, T., Lee, G., Lazic, A. et al. Inhibition of a viral enzyme by a small-molecule dimer disruptor. Nat Chem Biol 5, 640–646 (2009). https://doi.org/10.1038/nchembio.192

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