Laboratory and Basic Science Research Commons^™

A Biochemical Approach To Define The Interactome For Calpain2 In Endothelial Cells, Dorothy E. Croall Phd, Jordan R. Hoffman Ms, Calvin Ph Vary Phd, Sharon L. Ashworth Phd, Billy R. Chase, Jay Beaudoin Ii

Molecular & Biomedical Sciences Faculty Scholarship

Current repositories for protein-protein interactions and high throughput screening methods focus on individual gene products and do not consider the significance of calcium induced conformational changes. These limitations suggest the need for alternative strategies to better define the calpain2 interactome. Affinity capture coupled with LC-MS/MS and proteomic analysis of the recovered proteins provides a powerful approach to identify protein-protein interactions for the heterodimeric calpain2. CAPN2 (rat) was modified to be catalytically incompetent (C105A) and fused with a C-terminal 15 residue peptide optimized for biotinylation by the biotin protein ligase, BirA. The resulting CAPN2*, heterodimerized with truncated CAPNS1, was purified from …