6oet is a 10 chain structure with sequence from Lk3 transgenic mice. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance.
[RAG1_MOUSE] Catalytic component of the RAG complex, a multiprotein complex that mediates the DNA cleavage phase during V(D)J recombination. V(D)J recombination assembles a diverse repertoire of immunoglobulin and T-cell receptor genes in developing B and T-lymphocytes through rearrangement of different V (variable), in some cases D (diversity), and J (joining) gene segments. In the RAG complex, RAG1 mediates the DNA-binding to the conserved recombination signal sequences (RSS) and catalyzes the DNA cleavage activities by introducing a double-strand break between the RSS and the adjacent coding segment. RAG2 is not a catalytic component but is required for all known catalytic activities. DNA cleavage occurs in 2 steps: a first nick is introduced in the top strand immediately upstream of the heptamer, generating a 3'-hydroxyl group that can attack the phosphodiester bond on the opposite strand in a direct transesterification reaction, thereby creating 4 DNA ends: 2 hairpin coding ends and 2 blunt, 5'-phosphorylated ends. The chromatin structure plays an essential role in the V(D)J recombination reactions and the presence of histone H3 trimethylated at 'Lys-4' (H3K4me3) stimulates both the nicking and haipinning steps. The RAG complex also plays a role in pre-B cell allelic exclusion, a process leading to expression of a single immunoglobulin heavy chain allele to enforce clonality and monospecific recognition by the B-cell antigen receptor (BCR) expressed on individual B-lymphocytes. The introduction of DNA breaks by the RAG complex on one immunoglobulin allele induces ATM-dependent repositioning of the other allele to pericentromeric heterochromatin, preventing accessibility to the RAG complex and recombination of the second allele. In addition to its endonuclease activity, RAG1 also acts as a E3 ubiquitin-protein ligase that mediates monoubiquitination of histone H3. Histone H3 monoubiquitination is required for the joining step of V(D)J recombination. Mediates polyubiquitination of KPNA1.[1][2][3][4][5][6][7][8][9][10][11][12][13] [RAG2_MOUSE] Core component of the RAG complex, a multiprotein complex that mediates the DNA cleavage phase during V(D)J recombination. V(D)J recombination assembles a diverse repertoire of immunoglobulin and T-cell receptor genes in developing B and T-lymphocytes through rearrangement of different V (variable), in some cases D (diversity), and J (joining) gene segments. DNA cleavage by the RAG complex occurs in 2 steps: a first nick is introduced in the top strand immediately upstream of the heptamer, generating a 3'-hydroxyl group that can attack the phosphodiester bond on the opposite strand in a direct transesterification reaction, thereby creating 4 DNA ends: 2 hairpin coding ends and 2 blunt, 5'-phosphorylated ends. The chromatin structure plays an essential role in the V(D)J recombination reactions and the presence of histone H3 trimethylated at 'Lys-4' (H3K4me3) stimulates both the nicking and haipinning steps. The RAG complex also plays a role in pre-B cell allelic exclusion, a process leading to expression of a single immunoglobulin heavy chain allele to enforce clonality and monospecific recognition by the B-cell antigen receptor (BCR) expressed on individual B-lymphocytes. The introduction of DNA breaks by the RAG complex on one immunoglobulin allele induces ATM-dependent repositioning of the other allele to pericentromeric heterochromatin, preventing accessibility to the RAG complex and recombination of the second allele. In the RAG complex, RAG2 is not the catalytic component but is required for all known catalytic activities mediated by RAG1. It probably acts as a sensor of chromatin state that recruits the RAG complex to H3K4me3.[14][15][16][17][18][19]
Publication Abstract from PubMed
The RAG1-RAG2 recombinase (RAG) cleaves DNA to initiate V(D)J recombination, but RAG also belongs to the RNH-type transposase family. To learn how RAG-catalyzed transposition is inhibited in developing lymphocytes, we determined the structure of a DNA-strand transfer complex of mouse RAG at 3.1-A resolution. The target DNA is a T form (T for transpositional target), which contains two >80 degrees kinks towards the minor groove, only 3 bp apart. RAG2, a late evolutionary addition in V(D)J recombination, appears to enforce the sharp kinks and additional inter-segment twisting in target DNA and thus attenuates unwanted transposition. In contrast to strand transfer complexes of genuine transposases, where severe kinks occur at the integration sites of target DNA and thus prevent the reverse reaction, the sharp kink with RAG is 1 bp away from the integration site. As a result, RAG efficiently catalyzes the disintegration reaction that restores the RSS (donor) and target DNA.
How mouse RAG recombinase avoids DNA transposition.,Chen X, Cui Y, Wang H, Zhou ZH, Gellert M, Yang W Nat Struct Mol Biol. 2020 Feb;27(2):127-133. doi: 10.1038/s41594-019-0366-z. Epub, 2020 Feb 3. PMID:32015553[20]
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.
References
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↑ McBlane JF, van Gent DC, Ramsden DA, Romeo C, Cuomo CA, Gellert M, Oettinger MA. Cleavage at a V(D)J recombination signal requires only RAG1 and RAG2 proteins and occurs in two steps. Cell. 1995 Nov 3;83(3):387-95. PMID:8521468
↑ Agrawal A, Schatz DG. RAG1 and RAG2 form a stable postcleavage synaptic complex with DNA containing signal ends in V(D)J recombination. Cell. 1997 Apr 4;89(1):43-53. PMID:9094713
↑ Landree MA, Wibbenmeyer JA, Roth DB. Mutational analysis of RAG1 and RAG2 identifies three catalytic amino acids in RAG1 critical for both cleavage steps of V(D)J recombination. Genes Dev. 1999 Dec 1;13(23):3059-69. PMID:10601032
↑ Fugmann SD, Villey IJ, Ptaszek LM, Schatz DG. Identification of two catalytic residues in RAG1 that define a single active site within the RAG1/RAG2 protein complex. Mol Cell. 2000 Jan;5(1):97-107. PMID:10678172
↑ Jones JM, Gellert M. Autoubiquitylation of the V(D)J recombinase protein RAG1. Proc Natl Acad Sci U S A. 2003 Dec 23;100(26):15446-51. Epub 2003 Dec 11. PMID:14671314 doi:http://dx.doi.org/10.1073/pnas.2637012100
↑ Lu CP, Sandoval H, Brandt VL, Rice PA, Roth DB. Amino acid residues in Rag1 crucial for DNA hairpin formation. Nat Struct Mol Biol. 2006 Nov;13(11):1010-5. Epub 2006 Oct 8. PMID:17028591 doi:http://dx.doi.org/10.1038/nsmb1154
↑ Shimazaki N, Tsai AG, Lieber MR. H3K4me3 stimulates the V(D)J RAG complex for both nicking and hairpinning in trans in addition to tethering in cis: implications for translocations. Mol Cell. 2009 Jun 12;34(5):535-44. doi: 10.1016/j.molcel.2009.05.011. PMID:19524534 doi:http://dx.doi.org/10.1016/j.molcel.2009.05.011
↑ Simkus C, Makiya M, Jones JM. Karyopherin alpha 1 is a putative substrate of the RAG1 ubiquitin ligase. Mol Immunol. 2009 Apr;46(7):1319-25. doi: 10.1016/j.molimm.2008.11.009. Epub 2008, Dec 31. PMID:19118899 doi:http://dx.doi.org/10.1016/j.molimm.2008.11.009
↑ Hewitt SL, Yin B, Ji Y, Chaumeil J, Marszalek K, Tenthorey J, Salvagiotto G, Steinel N, Ramsey LB, Ghysdael J, Farrar MA, Sleckman BP, Schatz DG, Busslinger M, Bassing CH, Skok JA. RAG-1 and ATM coordinate monoallelic recombination and nuclear positioning of immunoglobulin loci. Nat Immunol. 2009 Jun;10(6):655-64. doi: 10.1038/ni.1735. PMID:19448632 doi:http://dx.doi.org/10.1038/ni.1735
↑ Grazini U, Zanardi F, Citterio E, Casola S, Goding CR, McBlane F. The RING domain of RAG1 ubiquitylates histone H3: a novel activity in chromatin-mediated regulation of V(D)J joining. Mol Cell. 2010 Jan 29;37(2):282-93. doi: 10.1016/j.molcel.2009.12.035. PMID:20122409 doi:http://dx.doi.org/10.1016/j.molcel.2009.12.035
↑ Yin FF, Bailey S, Innis CA, Ciubotaru M, Kamtekar S, Steitz TA, Schatz DG. Structure of the RAG1 nonamer binding domain with DNA reveals a dimer that mediates DNA synapsis. Nat Struct Mol Biol. 2009 May;16(5):499-508. Epub 2009 Apr 26. PMID:19396172 doi:http://dx.doi.org/10.1038/nsmb.1593
↑ Oettinger MA, Schatz DG, Gorka C, Baltimore D. RAG-1 and RAG-2, adjacent genes that synergistically activate V(D)J recombination. Science. 1990 Jun 22;248(4962):1517-23. PMID:2360047
↑ McBlane JF, van Gent DC, Ramsden DA, Romeo C, Cuomo CA, Gellert M, Oettinger MA. Cleavage at a V(D)J recombination signal requires only RAG1 and RAG2 proteins and occurs in two steps. Cell. 1995 Nov 3;83(3):387-95. PMID:8521468
↑ Agrawal A, Schatz DG. RAG1 and RAG2 form a stable postcleavage synaptic complex with DNA containing signal ends in V(D)J recombination. Cell. 1997 Apr 4;89(1):43-53. PMID:9094713
↑ West KL, Singha NC, De Ioannes P, Lacomis L, Erdjument-Bromage H, Tempst P, Cortes P. A direct interaction between the RAG2 C terminus and the core histones is required for efficient V(D)J recombination. Immunity. 2005 Aug;23(2):203-12. PMID:16111638 doi:http://dx.doi.org/10.1016/j.immuni.2005.07.004
↑ Shimazaki N, Tsai AG, Lieber MR. H3K4me3 stimulates the V(D)J RAG complex for both nicking and hairpinning in trans in addition to tethering in cis: implications for translocations. Mol Cell. 2009 Jun 12;34(5):535-44. doi: 10.1016/j.molcel.2009.05.011. PMID:19524534 doi:http://dx.doi.org/10.1016/j.molcel.2009.05.011
↑ Hewitt SL, Yin B, Ji Y, Chaumeil J, Marszalek K, Tenthorey J, Salvagiotto G, Steinel N, Ramsey LB, Ghysdael J, Farrar MA, Sleckman BP, Schatz DG, Busslinger M, Bassing CH, Skok JA. RAG-1 and ATM coordinate monoallelic recombination and nuclear positioning of immunoglobulin loci. Nat Immunol. 2009 Jun;10(6):655-64. doi: 10.1038/ni.1735. PMID:19448632 doi:http://dx.doi.org/10.1038/ni.1735
↑ Chen X, Cui Y, Wang H, Zhou ZH, Gellert M, Yang W. How mouse RAG recombinase avoids DNA transposition. Nat Struct Mol Biol. 2020 Feb;27(2):127-133. doi: 10.1038/s41594-019-0366-z. Epub, 2020 Feb 3. PMID:32015553 doi:http://dx.doi.org/10.1038/s41594-019-0366-z
