Crystallographic Studies On Recombinant PX Domain Of SNX11

Sorting Nexins (SNXs) are a large group of proteins defined by the presence of a conserved Phox homology (PX) domain and their critical roles in regulating intracellular membrane trafficking. Sorting nexin 11 (SNX11) was identified, by searching human expressed sequence tag databases with human SNX1, as a member of the SNX family. Both sequence alignment with other SNXs and secondary structure prediction revealed that SNX11 contains the Phox homology (PX) domain which is conserved in all members of this family. While most SNXs within the subgroup that SNX11 belongs to have been shown to play critical roles in protein trafficking, none have been reported for SNX11. Structural study is necessary to gain insight for the function of SNX11.Here, we report the crystallization and preliminary X-ray diffraction analysis of the PX domain of human Sorting Nexin (hSNX-PX) and mouse Sorting Nexin 11 (mSNX11-PX). Three C-terminal His-tagged truncated versions contained PX domain of hSNX11 and one mSNX11-PX were successfully overexpressed in a soluble form in E. coli. Gel-filtration purification analysis showed that all four of the truncated SNX11 were monomeric in solution. The purities were higher than 95% as determined by SDS-PAGE. Crystals were obtained using the sitting-drop vapor-diffusion method at 293 K. After crystallization optimization, all of the truncated versions had high diffraction quality for structural solution. We have collected the diffraction data of the three truncated versions of hSNX-PX from Shanghai Synchrotron Radiation Facility and mSNX11-PX using an in-house X-ray source at 100 K. The four structures have been solved by molecular replacement. Further model improvement and refinement are in progress.