Crystallization and preliminary diffraction analysis of DUSP28 through identification of a pseudo-thrombin cleavage site

Abstract

Dual specificity protein phosphatases (DUSPs) belong to the protein tyrosine phosphatase (PTP) family. DUSPsdephosphorylate both phospho-serine/threonine and phospho-tyrosine of mitogen activated protein kinases (MAPKs)and play important roles in cell growth, regulation and signaling. DUSP28, a member of the atypical DUSPs, hasdephosphorylation activity towards proteins involved in cellular signaling processes. DUSP28 is also implicated in thedevelopment of pancreatic cancer and liver cancer. The atomic resolution structure of DUSP28 should help the structurebaseddesign of specific and potent therapeutics. However, the structure and detailed function of DUSP28 have not beenelucidated yet. Here, we prepared a large quantity of DUSP28 protein and crystallized the protein. During the proteinpreparation, we encountered an unexpected proteolytic cleavage in the middle of the protein domain and overcamethe problem by identifying and mutating the pseudo-thrombin cleavage site. By using the purified protein, we were ableto grow diffraction quality crystals and collected a 2.1 A resolution diffraction data. The preliminary diffraction analysisrevealed that the crystal is in the space group P3121 with unit cell parameters of a = 78.85 A, b = 78.85 A, c = 90.26 A, α =90.00°, β = 90.00° and γ = 120.00°