Summary
V(D)J recombination in the vertebrate immune system generates a highly diverse population of immunoglobulins and T cell receptors by combinatorial joining of segments of coding DNA. The RAG1-RAG2 protein complex (RAG1/2) initiates this site-specific recombination by cutting DNA at specific sites flanking the coding segments. We report here the crystal structure of the RAG1/2 complex at 3.2Å resolution. The 230 kDa RAG1/2 heterotetramer is Y-shaped, with the N-terminal domains of the two RAG1 chains forming an intertwined stalk. Each RAG1/2 heterodimer composes one arm of the “Y” with the active site in the center and RAG2 at its tip. The RAG1/2 structure rationalizes more than 60 mutations identified in immunodeficient patients, as well as a large body of genetic and biochemical data. The architectural similarity between RAG1 and the hairpin-forming transposases Hermes and Tn5 suggests the evolutionary conservation of these DNA rearrangements.
Structures of type-1 human immunodeficiency virus (HIV-1) reverse transcriptase (RT) have been determined in several forms, but only one contains an RNA/DNA hybrid. Here we report three structures of HIV-1 RT complexed with a non-nucleotide RT inhibitor (NNRTI) and an RNA/DNA hybrid. In the presence of an NNRTI, the RNA/DNA structure differs from all prior nucleic acid bound to RT including the RNA/DNA hybrid. The enzyme structure also differs from all previous RT–DNA complexes. As a result, the hybrid has ready access to the RNase H active site. These observations indicate that an RT–nucleic acid complex may adopt two structural states, one competent for DNA polymerization and the other for RNA degradation. RT mutations that confer drug resistance but are distant from the inhibitor-binding sites often map to the unique RT–hybrid interface that undergoes conformational changes between two catalytic states.
Type I restriction-modification enzymes act as conventional adenine methylases on hemimethylated DNAs, but unmethylated recognition targets induce them to translocate thousands of base pairs before cleaving distant sites nonspecifically. The first crystal structure of a type I motor subunit responsible for translocation and cleavage suggests how the pentameric translocating complex is assembled and provides a structural framework for translocation of duplex DNA by RecA-like ATPase motors.
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