Abstract
Maintaining the biological functionality of immobilized proteins is the key to the success of numerous protein-based biomedical devices. To that end, we studied conformational change of calmodulin (CaM) immobilized on chemical patterns. 1-cysteine mutated calmodulin was immobilized on a mercapto-terminated surface through the cysteine-Hg-mercapto coupling. Utilizing Atomic Force Microscope (AFM), the average height of the immobilized calmodulin was determined to be 1.87 ± 0.19 nm. After incubation in EGTA solution, the average height of protein changed to 2.26 ± 0.21 nm, indicating conformational change of CaM to Apo-CaM. The immobilized CaM also demonstrated conformational change upon the reaction with known calmodulin antagonist chlorpromazine (CPZ). After incubation in CPZ solution, the average height of CPZ-bound CaM increased to 2.32 ± 0.20 nm, demonstrating the immobilized CaM still has the similar response as in bulk solution. These results show that immobilization of calmodulin on a solid support does not interfere with the ability of the protein to bind calcium and calmodulin antagonists. Our results demonstrate the feasibility of employing AFM to probe and understand protein conformational changes.
