Computational protocol: X-ray structure of a carpet-like antimicrobial defensin–phospholipid membrane disruption complex

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Protocol publication

[…] The NaD1–PA complexes were generated by mixing NaD1 with 08:0 PA (Avanti Polar Lipids) to a protein:lipid molar ratio of 1:2 (8 mg/mL:1.3 mg/mL). Crystals were grown over 5 days in sitting drops at 20 °C by mixing 150 nL protein–lipid solution with 150 nL well solution containing 28% PEG 4000, 1 M sodium chloride, and 0.1 M trisodium-citrate-citric acid pH 5.9. Crystals were flash-cooled at 100 K in mother liquor supplemented with 20% ethylene glycol, and data were collected at the Australian Synchrotron (MX2) and processed using XDS. The structure was solved by molecular replacement with PHASER using the structure of NaD1 in its dimeric form as a search model. The final model was built with Coot and refined with Phenix to a resolution of 2.5 Å with the final Rwork and Rfree values of 0.2031 and 0.2519, respectively. All data collection and refinement statistics are summarized in Table . All programs were accessed via the SBGrid suite. Figures were prepared using PyMol. Movies were prepared using Chimera. […]

Pipeline specifications

Software tools XDS, Coot, PHENIX, PyMOL
Applications Small-angle scattering, Protein structure analysis
Organisms Dipturus trachyderma, Fungi
Chemicals Phosphatidic Acids