Purification, crystallization and X-ray crystallographic analysis of RPTPH

Abstract

Receptor-type protein tyrosine phosphatases (RPTPs) belong to the protein tyrosine phosphatase (PTP) family and play a critical role in cell signaling. RPTPH, a type of RPTP, consists of long extracellular domains, a transmembrane domain, and a single intracellular domain with phosphatase activity. RPTPH is involved in phosphorylation of target proteins involved in the AKT signaling pathway and regulates T-cell function and cell apoptosis. The protein is also implicated in progression of colorectal and lung cancers. Despite the importance of RPTPH in tumor-related cell signaling and therapeutic drug development, the structure of this enzyme has not yet been determined. We overexpressed, purified, and crystallized the catalytic domain of RPTPH. The RPTPH crystal diffracted at a resolution of 1.56 Å. It belonged to the space group P3 2 with unit cell parameters a = b = 56.46 Å, c = 80.45 Å, α = β = 90°, and γ = 120°.

Crystal structure of the catalytic domain of human RPTPH

“…Two mutations, C818A and C1020S, were introduced to prevent oxidation of the protein. Detailed information on protein expression, purification and crystallization has previously been reported (Kim & Ryu, 2021). Briefly, Escherichia coli BL21(DE3) cells transformed with the pET-28a vector encoding the mutated RPTPH were cultured in Luria-Bertani medium and protein overexpression was induced with isopropyl -d-1-thiogalactopyranoside.…”