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Second harmonic generation detection of Ras conformational changes and discovery of a small molecule binder

  1. Author:
    Donohue, Elizabeth
    Khorsand, Sina
    Mercado, Gabriel
    Varney, Kristen M
    Wilder, Paul T
    Yu, Wenbo
    MacKerell, Alexander D
    Alexander,Patrick
    Van,Que
    Moree, Ben [ORCID]
    Stephen,Andy
    Weber, David J [ORCID]
    Salafsky, Joshua
    McCormick, Frank

  2. Author Address

    Helen Diller Family Comprehensive Cancer Center, University of California, San Francisco, CA 94158., Biodesy, Inc., South San Francisco, CA 94080., Center for Biomolecular Therapeutics, School of Medicine, University of Maryland, Baltimore, MD 21201., Department of Biochemistry and Molecular Biology, School of Medicine, University of Maryland, Baltimore, MD 21201., Marlene and Stewart Greenebaum Comprehensive Cancer Center, University of Maryland, Baltimore, MD 21201., Department of Pharmaceutical Sciences, School of Pharmacy, University of Maryland, Baltimore, MD 21201., National Cancer Institute RAS Initiative, Cancer Research Technology Program, Frederick National Laboratory for Cancer Research, Leidos Biomedical Research, Inc., Frederick, MD 21702., Department of Pharmaceutical Chemistry, University of California, San Francisco, CA 94158 frank.mccormick@ucsf.edu joshua.salafsky@ucsf.edu., Helen Diller Family Comprehensive Cancer Center, University of California, San Francisco, CA 94158; frank.mccormick@ucsf.edu joshua.salafsky@ucsf.edu.,
    1. Year: 2019
    2. Date: Aug 27
    3. Epub Date: 2019 08 09
  2. Journal: Proceedings of the National Academy of Sciences of the United States of America
    1. 116
    2. 35
    3. Pages: 17290-17297
  4. Type of Article: Article
  5. ISSN: 0027-8424
  1. Abstract:

    Second harmonic generation (SHG) is an emergent biophysical method that sensitively measures real-time conformational change of biomolecules in the presence of biological ligands and small molecules. This study describes the successful implementation of SHG as a primary screening platform to identify fragment ligands to oncogenic Kirsten rat sarcoma (KRas). KRas is the most frequently mutated driver of pancreatic, colon, and lung cancers; however, there are few well-characterized small molecule ligands due to a lack of deep binding pockets. Using SHG, we identified a fragment binder to KRasG12D and used 1H 15N transverse relaxation optimized spectroscopy (TROSY) heteronuclear single-quantum coherence (HSQC) NMR to characterize its binding site as a pocket adjacent to the switch 2 region. The unique sensitivity of SHG furthered our study by revealing distinct conformations induced by our hit fragment compared with 4,6-dichloro-2-methyl-3-aminoethyl-indole (DCAI), a Ras ligand previously described to bind the same pocket. This study highlights SHG as a high-throughput screening platform that reveals structural insights in addition to ligand binding.

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External Sources

  1. View Full Text
  2. DOI: 10.1073/pnas.1905516116
  3. PMID: 31399543
  4. View record in Web of Science®
  5. WOS: 000483396800028
  6. PII : 1905516116

Library Notes

  1. Fiscal Year: FY2018-2019
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