Tzu-Ching Meng
Academia Sinica, Taiwan
Title: Structural analysis of PTPN3-p38y interactions by chemical cross-linking coupled with mass spectrometry
Biography
Biography: Tzu-Ching Meng
Abstract
Purpose: Protein tyrosine phosphatase N3 (PTPN3) and mitogen-activated protein kinase p38y coordinae to promote Ras-induced oncegenesis. Structural analysis of PTPN3-p38y complex is a challenging task by using traditional x-ray crystallography. Therefore, chemical cross-linking coupled with mass spectrometry (CX-MS) has been developed as an alternative method to solve this obstacle. Experimental description: Through covalent linkage of two spatially proximate residues within a single or between two polypeptide chains, the CX-MS approach provides structural insights into the flexible regions of proteins under any circumstances amenable to analysis. Conventionally, mapping of the protein-protein interaction sites relies on the coupling of lysine residues on the surface. Together with the distance restraint of the selected cross-linker, the segments of interaction surface can be determined. Results: Employing this method, we have mapped the contact interfaces of the PTPN3-p38y complex. Our finding of the solution structure indicated that the catalytic domain of PTPN3 interacts with the activation loop of phosphorylated p38y, illustrating how PTPN3-mediated dephosphorylation of p38y takes place. The CX-MS approach further demonstrated that the PDZ domain of PTPN3 recruits the PDZ-binding motif of p38y, thus stabilizing the active-state complex of PTPN3-p38y. Moreover, the CX-MS analysis defined the autoinhibitory characteristic of the PDZ domain in PTPN3 in the absence of p38y. Conclusions: By combining other approaches such as small-angle x-ray scattering (SAXA) and crystal structure of PTPN3-phospho-p38y peptide, the CX-MS method generates considerable insights into the architecture of the phosphatase-kinase complex assembly.
