Anti-CRISPR proteins function through thermodynamic tuning and allosteric regulation of CRISPR RNA-guided surveillance complex
- Patterson, Angela [ Montana State University: Chemistry & Biochemistry ]
- White, Aidan [ Montana State University: Chemistry & Biochemistry ]
- Waymire, Elizabeth [ Montana State University: Chemistry & Biochemistry ]
- Fleck, Sophie [ Montana State University: Chemistry & Biochemistry ]
- Golden, Sarah [ Montana State University: Microbiology & Cell Biology ]
- Wilkinson, Royce A. [ Montana State University: Microbiology & Cell Biology ]
- Wiedenheft, Blake [ Montana State University: Microbiology & Cell Biology ]
- Bothner, Brian [ Montana State University: Microbiology & Cell Biology ]
CRISPR RNA-guided detection and degradation of foreign DNA is a dynamic process. Viruses can interfere with this cellular defense by expressing small proteins called anti-CRISPRs. While structural models of anti-CRISPRs bound to their target complex provide static snapshots that inform mechanism, the dynamics and thermodynamics of these interactions are often overlooked. Here, we use hydrogen deuterium exchange-mass spectrometry (HDX-MS) and differential scanning fluorimetry (DSF) experiments to determine how anti-CRISPR binding impacts the conformational landscape of the type IF CRISPR RNA guided surveillance complex (Csy) upon binding of two different anti-CRISPR proteins (AcrIF9 and AcrIF2). The results demonstrate that AcrIF2 binding relies on enthalpic stabilization, whereas AcrIF9 uses an entropy driven reaction to bind the CRISPR RNA-guided surveillance complex. Collectively, this work reveals the thermodynamic basis and mechanistic versatility of anti-CRISPR-mediated immune suppression. More broadly, this work presents a striking example of how allosteric effectors are employed to regulate nucleoprotein complexes.