PDBsum entry 6dbr

Recombination/DNA

PDB id: 6dbr

Contents

Protein chains

550 a.a.

351 a.a.

DNA/RNA

Metals

_CA ×4

_ZN ×2

PDB id:

6dbr

Name:

Recombination/DNA

Title:

Cryo-em structure of rag in complex with one melted rss and one unmelted rss

Structure:

Recombination activating gene 1 - mbp chimera. Chain: a, c. Synonym: mbp,mmbp,maltodextrin-binding protein,rag-1. Engineered: yes. Recombination activating gene 2. Chain: b, d. Synonym: v(d)j recombination-activating protein 2. Engineered: yes. Forward strand of unmelted rss substrate DNA.

Source:

Escherichia coli, danio rerio. Zebrafish. Organism_taxid: 83333, 7955. Strain: k12. Gene: male, b4034, jw3994, rag1. Expressed in: spodoptera frugiperda. Expression_system_taxid: 7108. Danio rerio. Organism_taxid: 7955.

Authors:

H.Wu,M.Liao,H.Ru,W.Mi

Key ref:

H.Ru et al. (2018). DNA melting initiates the RAG catalytic pathway. Nat Struct Mol Biol, 25, 732-742. PubMed id: 30061602

Date:

03-May-18 Release date: 01-Aug-18

Protein chains

O13033  (RAG1_DANRE) -  V(D)J recombination-activating protein 1 from Danio rerio

Seq: 1057 a.a.

Struc: 550 a.a.

Protein chains

P0AEX9  (MALE_ECOLI) -  Maltose/maltodextrin-binding periplasmic protein from Escherichia coli (strain K12)

Seq: 396 a.a.

Struc: 550 a.a.*

Protein chains

O13034  (RAG2_DANRE) -  V(D)J recombination-activating protein 2 from Danio rerio

Seq: 530 a.a.

Struc: 351 a.a.*

* PDB and UniProt seqs differ at 312 residue positions (black crosses)

DNA/RNA chains

G-A-T-C-T-G-G-C-C-T-G-T-C-T-T-A-C-A-C-A-G-T-G-C-T-A-C-A-G-A-C-T-G-G 34 bases

C-C-A-G-T-C-T-G-T-A-G-C-A-C-T-G-T-G-T-A-A-G-A-C-A-G-G-C-C-A-G-A-T-C 34 bases

G-A-T-C-T-G-G-C-C-T-G-T-C-T-T-A-C-A-C-A-G-T-G-G-T-A-G-T-A-C-T-C-C-A 34 bases

T-G-G-A-G-T-A-C-T-A-C-C-A-C-T-G-T-G-T-A-A-G-A-C-A-G-G-C-C-A-G-A-T-C 34 bases

Enzyme reactions

• Enzyme class 2: Chains A, C: E.C.2.3.2.27 - RING-type E3 ubiquitin transferase.
• Reaction: S-ubiquitinyl-[E2 ubiquitin-conjugating enzyme]-L-cysteine + [acceptor protein]-L-lysine = [E2 ubiquitin-conjugating enzyme]-L-cysteine + N₆- ubiquitinyl-[acceptor protein]-L-lysine
• Enzyme class 3: Chains A, C: E.C.3.1.-.-

Note, where more than one E.C. class is given (as above), each may correspond to a different protein domain or, in the case of polyprotein precursors, to a different mature protein.

Nat Struct Mol Biol 25:732-742 (2018)

PubMed id: 30061602

DNA melting initiates the RAG catalytic pathway.

H.Ru, W.Mi, P.Zhang, F.W.Alt, D.G.Schatz, M.Liao, H.Wu.

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 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.