Modeling of the RAG reaction mechanism.
In vertebrate V(D)J recombination, it remains unclear how the RAG complex coordinates its catalytic steps with binding to two distant recombination sites. Here, we test the ability of the plausible reaction schemes to fit observed time courses for RAG nicking and DNA hairpin formation. The reaction schemes with the best fitting capability (1) strongly favor a RAG tetrameric complex over a RAG octameric complex; (2) indicate that once a RAG complex brings two recombination signal sequence (RSS) sites into synapsis, the synaptic complex rarely disassembles; (3) predict that the binding of both RSS sites (synapsis) occurs before catalysis (nicking); and (4) show that the RAG binding properties permit strong distinction between RSS sites within active chromatin versus nonspecific DNA or RSS sites within inactive chromatin. The results provide general insights for synapsis by nuclear proteins as well as more specific testable predictions for the RAG proteins.
USC Norris Comprehensive Cancer Center, University of Southern California Keck School of Medicine, 1441 Eastlake Avenue, Los Angeles, CA 90089-9176, USA.
Amjad
A
Askary
USC Norris Comprehensive Cancer Center, University of Southern California Keck School of Medicine, 1441 Eastlake Avenue, Los Angeles, CA 90089-9176, USA.
Noriko
N
Shimazaki
Department of Chemical Engineering and Materials Science, University of Southern California, Los Angeles, CA 90089, USA.
Niki
N
Bayat
USC Norris Comprehensive Cancer Center, University of Southern California Keck School of Medicine, 1441 Eastlake Avenue, Los Angeles, CA 90089-9176, USA. Electronic address: lieber@usc.edu.
Michael R
MR
Lieber
D002843:Chromatin / Q000737:chemistry / Q000378:metabolism; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D018398:Homeodomain Proteins / Q000378:metabolism*; D006801:Humans; D055029:Inverted Repeat Sequences; D007700:Kinetics; D008956:Models, Chemical; D009687:Nuclear Proteins / Q000378:metabolism*; D011485:Protein Binding
D002843:Chromatin; D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; D009687:Nuclear Proteins; C499311:RAG2 protein, human; C064658:RAG-1 protein; D004247:DNA