Improved Electrophoretic Separation to Assist the Monitoring of Bcl-xL Post-Translational Modifications

Abstract

Bcl-x_L is an oncogene of which the survival functions are finely tuned by post-translational modifications (PTM). Within the Bcl-2 family of proteins, Bcl-x_L shows unique eligibility to deamidation, a time-related spontaneous reaction. Deamidation is still a largely overlooked PTM due to a lack of easy techniques to monitor Asn→Asp/IsoAsp conversions or Glu→Gln conversions. Being able to detect PTMs is essential to achieve a comprehensive description of all the regulatory mechanisms and functions a protein can carry out. Here, we report a gel composition improving the electrophoretic separation of deamidated forms of Bcl-x_L generated either by mutagenesis or by alkaline treatment. Importantly, this new gel formulation proved efficient to provide the long-sought evidence that even doubly-deamidated Bcl-x_L remains eligible for regulation by phosphorylation.