Expression, purification and crystallization of the SKICH domain of human TAX1BP1

Abstract

TAX1BP1 is a highly conserved, pleiotropic protein that plays many essential functions in human cells, including negative regulation of inflammatory and antimicrobial responses mediated by NF-κB and IRF3 signaling, inhibition of apoptosis, transcriptional coactivation and autophagy etc. TAX1BP1 contains a SKICH domain at the N-terminus, three coiled-coil domains in the middle and two ubiquitin-binding zinc-finger motifs at the C-terminus. The SKICH domain and the linker sequence between the SKICH domain and the coiled-coil region mediate interaction with ubiquitin-like proteins of the LC3/GABARAP family, which are autophagosome markers. For structure determination of the SKICH domain of TAX1BP1, a protein construct (amino acids 15-148) corresponding to the SKICH domain plus the linker region was expressed, purified and crystallized. A native diffraction data set has been collected to 1.9Å resolution. A molecular-replacement solution has been found by using the structure of the SKICH domain of NDP52, a paralog of TAX1BP1. © 2014 International Union of Crystallography All rights reserved.