Related Experiment Video
Updated: Aug 18, 2026

Preparation of the Mgm101 Recombination Protein by MBP-based Tagging Strategy
Published on: June 25, 2013
Determinants of HMGB proteins required to promote RAG1/2-recombination signal sequence complex assembly and catalysis
Yan Dai1, Ben Wong, Yi-Meng Yen
1Department of Molecular Biology, Massachusetts General Hospital, Boston, Massachusetts 02114, USA.
Abstract:
Efficient assembly of RAG1/2-recombination signal sequence (RSS) DNA complexes that are competent for V(D)J cleavage requires the presence of the nonspecific DNA binding and bending protein HMGB1 or HMGB2. We find that either of the two minimal DNA binding domains of HMGB1 is effective in assembling RAG1/2-RSS complexes on naked DNA and stimulating V(D)J cleavage but that both domains are required for efficient activity when the RSS is incorporated into a nucleosome. The single-domain HMGB protein from Saccharomyces cerevisiae, Nhp6A, efficiently assembles RAG1/2 complexes on naked DNA; however, these complexes are minimally competent for V(D)J cleavage. Nhp6A forms much more stable DNA complexes than HMGB1, and a variety of mutations that destabilize Nhp6A binding to bent microcircular DNA promote increased V(D)J cleavage. One of the two DNA bending wedges on Nhp6A and the analogous phenylalanine wedge at the DNA exit site of HMGB1 domain A were found to be essential for promoting RAG1/2-RSS complex formation. Because the phenylalanine wedge is required for specific recognition of DNA kinks, we propose that HMGB proteins facilitate RAG1/2-RSS interactions by recognizing a distorted DNA structure induced by RAG1/2 binding. The resulting complex must be sufficiently dynamic to enable the series of RAG1/2-mediated chemical reactions on the DNA.
More Related Videos
12:19Tools to Study the Role of Architectural Protein HMGB1 in the Processing of Helix Distorting, Site-specific DNA Interstrand Crosslinks
Published on: November 10, 2016
09:35Analysis of Somatic Hypermutation in the JH4 intron of Germinal Center B cells from Mouse Peyer's Patches
Published on: April 20, 2021
Related Concept Videos
Homologous Recombination
Conservative Site-specific Recombination and Phase Variation
The recognition sites for Cre recombinase called LoxP...
Crossing Over
Rab Proteins
Rab proteins switch between a cytosolic, GDP-bound inactive state and a membrane-anchored, GTP-bound active state. By themselves, Rabs show slow rates of GDP/GTP exchange and GTP hydrolysis. Thus, Rab proteins are considered...
Protein Complex Assembly
Many viruses self-assemble into a fully functional unit using the infected host cell to...
Restarting Stalled Replication Forks