Site-specific Identification and Quantification of Protein Glutathionylation under Ischemic Stress

Ischemia or ischemic reperfusion is well-known for its contribution to heart diseases. During ischemia and reperfusion, a flux of nutrients and oxygen to cardiomyocytes is altered, which causes a burst of ROS from mitochondria and other enzymes. Elevated levels of ROS can cause oxidative modifications of cardiac proteins, such as protein glutathionylation. Despite previous extensive studies, proteomic identification of glutathionylated proteins in cardiomyocytes under altered levels of nutrients and oxygen has been relatively limited. We have applied our clickable glutathione approach to HL-1 cardiomyocyte cell line under metabolic alterations of glucose, oxygen, and fatty acids to detect and identify proteins undergoing glutathionylation.