Computational protocol: Structure of TCR and antigen complexes at an immunodominant CTL epitope in HIV-1 infection

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Protocol publication

[…] All crystallizations were done by the sitting drop vapor diffusion method with a protein/reservoir drop ratio of 1:1 at 20°C. Crystallization conditions of grown crystals are shown in . For cryoprotection, obtained crystals were soaked briefly and sequentially in reservoir solutions containing 10% and 20% ethylene glycol, and then flash-frozen in liquid nitrogen. Data were collected at the beamline BL41XU in SPring 8 (Hyogo, Japan) and BL-5A, NW12A and BL1A in the PF facility (Tsukuba, Japan), and processed with HKL2000 and the CCP4 program suite. [...] The structures were determined by molecular replacement using Molrep. Search models used for molecular replacement were shown in . Model building and refinement were conducted using Coot and CNS 1.3, respectively. The further rounds of these refinements were performed using REFMAC implemented in CCP4. For building model of the T36-5 TCR-A24/Nef134-10(2F), diffraction intensities from the various crystals exhibited twin with an estimated twinning fraction of 0.45–0.49. Therefore, this structural model was refined using CNS 1.3 as a perfect twin.The stereochemistry of the refined models was assessed with program Rampage. All molecular graphics representations were created with the program PyMOL (DeLano Scientific; http://www.pymol.org). Data collection and refinement statistics are shown in . […]

Pipeline specifications

Software tools CCP4, Molrep, Coot, CNS, PyMOL
Application Protein structure analysis
Organisms Human immunodeficiency virus 1, Homo sapiens
Diseases HIV Infections
Chemicals Hydrogen