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Title | DNA melting initiates the RAG catalytic pathway. |
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Journal, issue, pages | Nat Struct Mol Biol, Vol. 25, Issue 8, Page 732-742, Year 2018 |
Publish date | Jul 30, 2018 |
Authors | Heng Ru / Wei Mi / Pengfei Zhang / Frederick W Alt / David G Schatz / Maofu Liao / Hao Wu / |
PubMed Abstract | The mechanism for initiating DNA cleavage by DDE-family enzymes, including the RAG endonuclease, which initiates V(D)J recombination, is not well understood. Here we report six cryo-EM structures of ...The mechanism for initiating DNA cleavage by DDE-family enzymes, including the RAG endonuclease, which initiates V(D)J recombination, is not well understood. Here we report six cryo-EM structures of zebrafish RAG in complex with one or two intact recombination signal sequences (RSSs), at up to 3.9-Å resolution. Unexpectedly, these structures reveal DNA melting at the heptamer of the RSSs, thus resulting in a corkscrew-like rotation of coding-flank DNA and the positioning of the scissile phosphate in the active site. Substrate binding is associated with dimer opening and a piston-like movement in RAG1, first outward to accommodate unmelted DNA and then inward to wedge melted DNA. These precleavage complexes show limited base-specific contacts of RAG at the conserved terminal CAC/GTG sequence of the heptamer, thus suggesting conservation based on a propensity to unwind. CA and TG overwhelmingly dominate terminal sequences in transposons and retrotransposons, thereby implicating a universal mechanism for DNA melting during the initiation of retroviral integration and DNA transposition. |
External links | Nat Struct Mol Biol / PubMed:30061602 / PubMed Central |
Methods | EM (single particle) |
Resolution | 3.0 - 5.0 Å |
Structure data | EMDB-7843, PDB-6dbi: EMDB-7844, PDB-6dbj: EMDB-7845, PDB-6dbl: EMDB-7846, PDB-6dbo: EMDB-7847, PDB-6dbq: EMDB-7848, PDB-6dbr: EMDB-7849, PDB-6dbt: EMDB-7850, PDB-6dbu: EMDB-7851, PDB-6dbv: |
Chemicals | ChemComp-ZN: ChemComp-CA: |
Source |
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Keywords | RECOMBINATION/DNA / V(D)J recombination / synaptic RAG complex / nicked RSS intermediates / paired complex / RECOMBINATION-DNA complex / RSS / RAG complex / melted DNA / Pre-cleavage complex / Pre-cleveage complex / RSS substrate DNA / Pre-cleveaage complex / Melted RSS / Unmelted RSS / unmelted DNA substrates / melted RSS substrate / singly-bound complex / unmelted RSS substrate / singly bound complex |