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Mol Cell. 2018 Apr 19;70(2):358-370.e4. doi: 10.1016/j.molcel.2018.03.008. Epub 2018 Apr 5.

Cracking the DNA Code for V(D)J Recombination.

Author information

1
Laboratory of Molecular Biology, NIDDK, NIH, Bethesda, MD 20892, USA; Integrative Bioscience and Biotechnology, Pohang University of Science and Technology, Pohang, Gyeongbuk 37673, Republic of Korea.
2
Laboratory of Molecular Biology, NIDDK, NIH, Bethesda, MD 20892, USA.
3
The California NanoSystems Institute, UCLA, Los Angeles, CA 90095, USA.
4
The California NanoSystems Institute, UCLA, Los Angeles, CA 90095, USA; Center of Cryo Electron Microscopy, Zhejiang University School of Medicine, Hangzhou, China.
5
Laboratory of Molecular Biology, NIDDK, NIH, Bethesda, MD 20892, USA. Electronic address: martinge@niddk.nih.gov.
6
Laboratory of Molecular Biology, NIDDK, NIH, Bethesda, MD 20892, USA. Electronic address: weiy@niddk.nih.gov.

Abstract

To initiate V(D)J recombination for generating the adaptive immune response of vertebrates, RAG1/2 recombinase cleaves DNA at a pair of recombination signal sequences, the 12- and 23-RSS. We have determined crystal and cryo-EM structures of RAG1/2 with DNA in the pre-reaction and hairpin-forming complexes up to 2.75 Å resolution. Both protein and DNA exhibit structural plasticity and undergo dramatic conformational changes. Coding-flank DNAs extensively rotate, shift, and deform for nicking and hairpin formation. Two intertwined RAG1 subunits crisscross four times between the asymmetric pair of severely bent 12/23-RSS DNAs. Location-sensitive bending of 60° and 150° in 12- and 23-RSS spacers, respectively, must occur for RAG1/2 to capture the nonamers and pair the heptamers for symmetric double-strand breakage. DNA pairing is thus sequence-context dependent and structure specific, which partly explains the "beyond 12/23" restriction. Finally, catalysis in crystallo reveals the process of DNA hairpin formation and its stabilization by interleaved base stacking.

KEYWORDS:

CA/TG dinucleotides; DNA bending; DNA hairpin; NBD; SCID; polypurine tract

PMID:
29628308
PMCID:
PMC5987255
DOI:
10.1016/j.molcel.2018.03.008
[Indexed for MEDLINE]
Free PMC Article

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