6P7M

Cryo-EM structure of LbCas12a-crRNA: AcrVA4 (1:2 complex)

Summary for 6P7M

• Entry DOI: 10.2210/pdb6p7m/pdb
• EMDB information: 20266 20267
• Descriptor: Cas12a, mature crRNA, anti-CRISPR VA4, ... (4 entities in total)
• Functional Keywords: crispr-cas, anti-crispr, cas12a, cpf1, lbcas12a, acrva4, rna binding protein-rna complex, rna binding protein/rna
• Biological source: Lachnospiraceae bacterium ND2006; More
• Total number of polymer chains: 3
• Total formula weight: 184666.27

Authors

Knott, G.J.,Liu, J.J.,Doudna, J.A. (deposition date: 2019-06-06, release date: 2019-08-21, Last modification date: 2024-03-20)

Primary citation

Knott, G.J.,Cress, B.F.,Liu, J.J.,Thornton, B.W.,Lew, R.J.,Al-Shayeb, B.,Rosenberg, D.J.,Hammel, M.,Adler, B.A.,Lobba, M.J.,Xu, M.,Arkin, A.P.,Fellmann, C.,Doudna, J.A.
Structural basis for AcrVA4 inhibition of specific CRISPR-Cas12a.
Elife, 8:-, 2019

PubMed Abstract: CRISPR-Cas systems provide bacteria and archaea with programmable immunity against mobile genetic elements. Evolutionary pressure by CRISPR-Cas has driven bacteriophage to evolve small protein inhibitors, anti-CRISPRs (Acrs), that block Cas enzyme function by wide-ranging mechanisms. We show here that the inhibitor AcrVA4 uses a previously undescribed strategy to recognize the Cas12a (LbCas12a) pre-crRNA processing nuclease, forming a Cas12a dimer, and allosterically inhibiting DNA binding. The Cas12a (AsCas12a) enzyme, widely used for genome editing applications, contains an ancestral helical bundle that blocks AcrVA4 binding and allows it to escape anti-CRISPR recognition. Using biochemical, microbiological, and human cell editing experiments, we show that Cas12a orthologs can be rendered either sensitive or resistant to AcrVA4 through rational structural engineering informed by evolution. Together, these findings explain a new mode of CRISPR-Cas inhibition and illustrate how structural variability in Cas effectors can drive opportunistic co-evolution of inhibitors by bacteriophage.

PubMed: 31397669
DOI: 10.7554/eLife.49110

Experimental method

ELECTRON MICROSCOPY (3 Å)