Abstract

Red blood cells (RBCs) transport oxygen and carbon dioxide as their main function and repeatedly circulate through the lung and capillaries during their 120 day life spans. RBCs concentrate large amounts of oxygen and are exposed continuously to intracellular reactive oxygen species (ROS) derived from autoxidation of oxyhemoglobin (oxyHb). Limiting the potential damage caused by these ROS is important in the RBC. Therefore, the antioxidant enzymes, CuZn-superoxide dismutase (CuZnSOD), catalase, glutathione peroxidase (GPx), and its substrate glutathione (GSH) are found in red blood cells. The localization of these antioxidants under physiological and oxidative stress conditions was examined. Our results showed that most CuZnSOD and catalase were concentrated at the RBC membrane and less in the cytosol, while GPx localized at the membrane under physiological conditions. After prolonged exposure of RBCs to air (1 h), CuZnSOD and catalase migrated to the membrane and they both associated with the cytoplamic domain of band 3 (CDB3). No association at the membrane of the three proteins was observed when RBCs were deaerated. Small amount of catalase and GPx localized to the membrane while CuZnSOD and GSH remained evenly distributed in the cytosol under two types (H 2 O 2 and O 2 - ) of ROS stress. In summary, CuZnSOD and catalase form a complex with CDB3 at the membrane, and the association of these proteins is mediated by the oxygenation state of the RBC. It is speculated that GPx cooperates with catalase to protect the whole RBC (membrane and cytoplasm) from ROS damage