Fluorescence resonance energy transfer analysis of recombination signal sequence configuration in the RAG1/2 synaptic complex.

Fluorescence resonance energy transfer analysis of recombination signal sequence configuration in the RAG1/2 synaptic complex. Research Abstract Details 

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Fluorescence resonance energy transfer analysis of recombination signal sequence configuration in the RAG1/2 synaptic complex. Abstract Text:

Mihai Ciubotaru,Aleksei N Kriatchko,Patrick C Swanson,Frank V Bright,David G Schatz,

A critical step in V(D)J recombination is the synapsis of complementary (12/23) recombination signal sequences (RSSs) by the RAG1 and RAG2 proteins to generate the paired complex (PC). Using a facilitated ligation assay and substrates that vary the helical phasing of the RSSs, we provide evidence that one particular geometric configuration of the RSSs is favored in the PC. To investigate this configuration further, we used fluorescent resonance energy transfer (FRET) to detect the synapsis of fluorescently labeled RSS oligonucleotides. FRET requires an appropriate 12/23 RSS pair, a divalent metal ion, and high-mobility-group protein HMGB1 or HMGB2. Energy transfer between the RSSs was detected with all 12/23 RSS end positions of the fluorescent probes but was not detected when probes were placed on the two ends of the same RSS. Energy transfer was confirmed to originate from the PC by using an in-gel FRET assay. The results argue against a unique planar configuration of the RSSs in the PC and are most easily accommodated by models in which synapsed 12- and 23-RSSs are bent and cross one another, with implications for the organization of the RAG proteins and the DNA substrates at the time of cleavage.

Fluorescence resonance energy transfer analysis of recombination signal sequence configuration in the RAG1/2 synaptic complex. Publishing Authors By Initials

M Ciubotaru,AN Kriatchko,PC Swanson,FV Bright,DG Schatz,

For similar genetic processes: recombination, genetic research abstracts see: genetic processes: recombination, genetic research

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Fluorescence resonance energy transfer analysis of recombination signal sequence configuration in the RAG1/2 synaptic complex. Journal Published:

PUBLICATION TYPE: Research Support, N.I.H., Extr

Journal: Molecular and cellular biology

VOLUME: 27

Page Numbers: 4745-58

Journal Abbreviation: Mol. Cell. Biol.

ISSN: 0270-7306

DAY: 30

MONTH: 04

YEAR: 2007

Fluorescence resonance energy transfer analysis of recombination signal sequence configuration in the RAG1/2 synaptic complex. Information

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LANGUAGE: eng

NlmUniqueID: 8109087

Fluorescence resonance energy transfer analysis of recombination signal sequence configuration in the RAG1/2 synaptic complex. Keywords Mesh Terms:

KEYWORDS: Recombination, Genetic

MESH TERMS: genetics

Chemical & Substance for Abstract: Fluorescence resonance energy transfer analysis of recombination signal sequence configuration in the RAG1/2 synaptic complex. Information

Substance Name: RAG-1 protein

Registry Number: 128559-51-3

Grant and Affiliation Information for Fluorescence resonance energy transfer analysis of recombination signal sequence configuration in the RAG1/2 synaptic complex.

AFFILIATION: Howard Hughes Medical Institute and Department of Immunibiology, Yale University School of Medicine, New Haven, CT 06520-8011, USA.

Country: United States

AGENCY: United States NIAID

GRANT: AI32524

ACRONYM: AI

MEDLINETA: Mol Cell Biol

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