Purification, crystallization and preliminary X-ray diffraction analysis of disease-related mutants of p97

The human type II Lambda Lambda Lambda+ protein p97 participates in various cellular activities, presumably through its involvement in the ubiquitin-proteasome degradation pathway. Mutations in p97 have been implicated in patients with inclusion-body myopathy associated with Paget's disease of the bone and frontotemporal dementia (IBMPFD). In this work, three mutant p97 N-D1 fragments, R86A, R95G and R155H, were crystallized in the presence of ATP gamma S with PEG 3350 as a main precipitant, yielding two different crystal forms. The R155H mutant crystal belonged to space group R3, with unit-cell parameters in the hexagonal setting of a = b = 134.2, c = 182.9 angstrom, and was merohedrally twinned, with an estimated twin fraction of 0.34. The crystals of the R86A and R95G mutants belonged to space group P1, with similar unit-cell parameters of a = 90.89, b = 102.6, c = 107.2 angstrom, alpha = 97.5, beta = 90.6, gamma = 91.5 degrees and a = 92.76, b = 103.7, c = 107.7 angstrom, alpha = 97.7, beta = 91.9, gamma = 89.7 degrees, respectively.