Use of High Pressure NMR Spectroscopy to Rapidly Identify Proteins with Internal Ligand-Binding Voids

Donald Gagné, Roksana Azad, Uthama R. Edupuganti, Justin Williams, James M. Aramini, Kazuyuki Akasaka, Kevin H. Gardner

Research output: Contribution to journalArticlepeer-review

Abstract

Small molecule binding within internal cavities provides a way to control protein function and structure, as exhibited in numerous natural and artificial settings. Unfortunately, most ways to identify suitable cavities require high-resolution structures a priori and may miss potential cryptic sites. Here we address this limitation via high-pressure solution NMR spectroscopy, taking advantage of the distinctive nonlinear pressure-induced chemical shift changes observed in proteins containing internal cavities and voids. We developed a method to rapidly characterize such nonlinearity among backbone 1H and 15N amide signals without needing to have sequence-specific chemical shift assignments, taking advantage of routinely available 15N-labeled samples, instrumentation, and 2D 1H/15N HSQC experiments. From such data, we find a strong correlation in the site-to-site variability in such nonlinearity with the total void volume within proteins, providing insights useful for prioritizing domains for ligand binding and indicating mode-of-action among such protein/ligand systems. We suggest that this approach provides a rapid and useful way to rapidly assess otherwise hidden dynamic architectures of protein that reflect fundamental properties associated with ligand binding and control.

Original languageEnglish (US)
JournalUnknown Journal
DOIs
StatePublished - Aug 26 2020

Keywords

  • High pressure NMR
  • Ligand binding
  • Protein cavities

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)
  • Immunology and Microbiology(all)
  • Neuroscience(all)
  • Pharmacology, Toxicology and Pharmaceutics(all)

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