Identification of SARS-CoV-2 S RBD escape mutants using yeast screening and deep mutational scanning

STAR Protoc. 2021 Dec 17;2(4):100869. doi: 10.1016/j.xpro.2021.100869. Epub 2021 Sep 22.

Abstract

Here, we describe a protocol to identify escape mutants on the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S) receptor-binding domain (RBD) using a yeast screen combined with deep mutational scanning. Over 90% of all potential single S RBD escape mutants can be identified for monoclonal antibodies that directly compete with angiotensin-converting enzyme 2 for binding. Six to 10 antibodies can be assessed in parallel. This approach has been shown to determine escape mutants that are consistent with more laborious SARS-CoV-2 pseudoneutralization assays. For complete details on the use and execution of this protocol, please refer to Francino-Urdaniz et al. (2021).

Keywords: Antibody; Immunology; Microbiology; Molecular Biology; Protein Biochemistry; Sequence analysis; Sequencing.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Angiotensin-Converting Enzyme 2 / genetics*
  • Angiotensin-Converting Enzyme 2 / metabolism
  • Binding Sites
  • COVID-19 / genetics*
  • COVID-19 / metabolism
  • COVID-19 / virology
  • DNA Mutational Analysis / methods*
  • Humans
  • Mutation*
  • SARS-CoV-2 / genetics*
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae / metabolism*
  • Spike Glycoprotein, Coronavirus / genetics*
  • Spike Glycoprotein, Coronavirus / metabolism

Substances

  • Spike Glycoprotein, Coronavirus
  • Angiotensin-Converting Enzyme 2