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Amplification, Next-generation Sequencing, and Genomic DNA Mapping of Retroviral Integration Sites
Published on: March 22, 2016
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Structural and sequencing analysis of local target DNA recognition by MLV integrase
Sriram Aiyer1, Paolo Rossi2, Nirav Malani3
1Department of Pharmacology, Robert Wood Johnson Medical School, Rutgers University, Piscataway, NJ 08854, USA.
Nucleic Acids Research
|May 14, 2015
Summary
Moloney murine leukemia virus integrase (IN) structural models reveal how IN binds target DNA. Specific regions of the IN catalytic core and C-terminal domains interact with DNA sequences near and far from integration sites.
Area of Science:
- Virology
- Structural Biology
- Molecular Biology
Background:
- Retroviral integrase (IN) proteins are crucial for viral pathogenesis, mediating target DNA integration.
- Understanding IN's target-site selection mechanism is key to controlling retroviral infections.
Purpose of the Study:
- To determine the structure of Moloney murine leukemia virus (M-MLV) IN domains.
- To elucidate the molecular interactions governing M-MLV IN target DNA binding and selection.
Main Methods:
- Solution nuclear magnetic resonance (NMR) spectroscopy for M-MLV IN C-terminal domain (CTD) structure.
- Homology modeling (SWISS-MODEL, MMM-tree, I-TASSER) for M-MLV IN catalytic core domain (CCD).
- In vivo mutagenesis, next-generation sequencing, and integration site analysis.
Main Results:
- The M-MLV IN CTD adopts an SH3 domain fold in solution.
- Structural models predicted DNA-binding residues in the CCD α2 helix and CTD β1-β2 loop.
- Mutational analysis confirmed CCD α2 helix (P187) interacts with distal DNA, and CTD β1-β2 loop with proximal DNA.
Conclusions:
- The study provides validated structural models of M-MLV IN domains.
- Identified specific IN-DNA interactions critical for retroviral integration target site selection.
- Findings advance understanding of retroviral DNA integration mechanisms.
