4QQZ

Crystal structure of T. fusca Cas3-AMPPNP

Summary for 4QQZ

• Entry DOI: 10.2210/pdb4qqz/pdb
• Related: 4QQX 4QQY 4QQZ
• Descriptor: CRISPR-associated helicase, Cas3 family, DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*A)-3'), FE (III) ION, ... (4 entities in total)
• Functional Keywords: hydrolase, helicase, hydrolase-dna complex, hydrolase/dna
• Biological source: Thermobifida fusca
• Total number of polymer chains: 8
• Total formula weight: 442168.08

Authors

Ke, A.,Huo, Y.,Nam, K.H. (deposition date: 2014-06-30, release date: 2014-08-27, Last modification date: 2024-02-28)

Primary citation

Huo, Y.,Nam, K.H.,Ding, F.,Lee, H.,Wu, L.,Xiao, Y.,Farchione, M.D.,Zhou, S.,Rajashankar, K.,Kurinov, I.,Zhang, R.,Ke, A.
Structures of CRISPR Cas3 offer mechanistic insights into Cascade-activated DNA unwinding and degradation.
Nat.Struct.Mol.Biol., 21:771-777, 2014

PubMed Abstract: CRISPR drives prokaryotic adaptation to invasive nucleic acids such as phages and plasmids, using an RNA-mediated interference mechanism. Interference in type I CRISPR-Cas systems requires a targeting Cascade complex and a degradation machine, Cas3, which contains both nuclease and helicase activities. Here we report the crystal structures of Thermobifida fusca Cas3 bound to single-stranded (ss) DNA substrate and show that it is an obligate 3'-to-5' ssDNase that preferentially accepts substrate directly from the helicase moiety. Conserved residues in the HD-type nuclease coordinate two irons for ssDNA cleavage. We demonstrate ATP coordination and conformational flexibility of the SF2-type helicase domain. Cas3 is specifically guided toward Cascade-bound target DNA by a PAM sequence, through physical interactions with both the nontarget substrate strand and the CasA protein. The sequence of recognition events ensures well-controlled DNA targeting and degradation of foreign DNA by Cascade and Cas3.

PubMed: 25132177
DOI: 10.1038/nsmb.2875

Experimental method

X-RAY DIFFRACTION (2.93 Å)