Preliminary X-ray crystallographic studies of mouse UPR responsive protein P58(IPK) TPR fragment

Abstract

Endoplasmic reticulum (ER) stress induces the unfolded protein response (UPR), which can promote protein folding and misfolded protein degradation and attenuate protein translation and protein translocation into the ER. P58(IPK) has been proposed to function as a molecular chaperone to maintain protein-folding homeostasis in the ER under normal and stressed conditions. P58(IPK) contains nine TPR motifs and a C-terminal J-domain within its primary sequence. To investigate the mechanism by which P58(IPK) functions to promote protein folding within the ER, a P58(IPK) TPR fragment without the C-terminal J-domain was crystallized. The crystals diffract to 2.5 A resolution using a synchrotron X-ray source. The crystals belong to space group P2(1), with unit-cell parameters a = 83.53, b = 92.75, c = 84.32 A, alpha = 90.00, beta = 119.36, gamma = 90.00 degrees. There are two P58(IPK) molecules in the asymmetric unit, which corresponds to a solvent content of approximately 60%. Structure determination by MAD methods is under way.

Figure 1

(a) A 13% SDS–PAGE gel of the purified mouse P58(IPK) fragment. The left lane contains protein standards of 97.4, 66, 45, 31, 21.5 and 14.5 kDa (from top to bottom). The right lane contains the purified mouse P58(IPK) fragment. (b) P58(IPK) fragment crystals. (c) Diffraction pattern of a P58(IPK) fragment crystal shown using the HKL-2000 package. The resolution at the detector edge is 2.50 Å.