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TitleStructural basis for AcrVA4 inhibition of specific CRISPR-Cas12a.
Journal, issue, pagesElife, Vol. 8, Year 2019
Publish dateAug 9, 2019
AuthorsGavin J Knott / Brady F Cress / Jun-Jie Liu / Brittney W Thornton / Rachel J Lew / Basem Al-Shayeb / Daniel J Rosenberg / Michal Hammel / Benjamin A Adler / Marco J Lobba / Michael Xu / Adam P Arkin / Christof Fellmann / Jennifer A Doudna /
PubMed AbstractCRISPR-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 ...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.
External linksElife / PubMed:31397669 / PubMed Central
MethodsEM (single particle)
Resolution3.0 - 4.9 Å
Structure data

20266

6p7m

EMDB-20266, PDB-6p7m:
Cryo-EM structure of LbCas12a-crRNA: AcrVA4 (1:2 complex)
Method: EM (single particle) / Resolution: 3.0 Å

20267

6p7n

EMDB-20267, PDB-6p7n:
Cryo-EM structure of LbCas12a-crRNA: AcrVA4 (2:2 complex)
Method: EM (single particle) / Resolution: 4.9 Å

Chemicals

ChemComp-MG:
Unknown entry

Source
  • lachnospiraceae bacterium nd2006 (bacteria)
  • moraxella bovoculi (bacteria)
KeywordsRNA BINDING PROTEIN/RNA / CRISPR-Cas / anti-CRISPR / Cas12a / Cpf1 / LbCas12a / AcrVA4 / RNA BINDING PROTEIN-RNA complex

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