In situ serial crystallography (iSX) structure of Autotaxin at room temperature

Measurements from rat ATX microcrystals grown in CrystalDirect plates were carried at room temperature (293 K) with a 4 × 4 μm2 microfocus beam at 14.2 keV (wavelength at 0.8731 Å) on ID23-2. Approximately 5.7 million diffraction frames from 61 drops across 2 CrystalDirect plates were collected using MeshScan, a modular automated ESRF workflow system (EWOKS) routine available through MXCuBE-Web. Each drop was raster scanned with a 4 ✕ 4 µm2 X-ray beam at 14.2 keV photon energy, a flux of 7x1011 ph/sec (i.e., 10% of full flux) and 6 ms exposure per frame without any rotation. This in-situ serial data collection strategy yielded ~100,000 (average) still diffraction frames per drop in ~10 min, ensuring the diffraction weighted dose to be ~35 kGy/diffraction frame as calculated using RADDOSE-3D. The still diffraction frames, collected at room temperature, were processed using crystFEL v.0.10.1. TRUNCATE from ccp4i was used to convert Intensities (I) to Structure Factors (SF). The structure was determined by molecular replacement using PHASER from phenix with the PDB model as a reference model. Refinement was carried out using phenix.refine for the reciprocal space and for real space.