Multiplexed silicon photonic sensor arrays enable facile characterization of coagulation protein binding to nanodiscs with variable lipid content.

ABSTRACT: Interactions of soluble proteins with the cell membrane are critical within the blood coagulation cascade. Of particular interest are the interactions of ?-carboxyglutamic acid-rich domain-containing clotting proteins with lipids. Variability among conventional analytical methods presents challenges for comparing clotting protein-lipid interactions. Most previous studies have investigated only a single clotting protein and lipid composition and have yielded widely different binding constants. Herein, we demonstrate that a combination of lipid bilayer nanodiscs and a multiplexed silicon photonic analysis technology enables high-throughput probing of many protein-lipid interactions among blood-clotting proteins. This approach allowed direct comparison of the binding constants of prothrombin, factor X, activated factor VII, and activated protein C to seven different binary lipid compositions. In a single experiment, the binding constants of one protein interacting with all lipid compositions were simultaneously determined. A simple surface regeneration then facilitated similar binding measurements for three other coagulation proteins. As expected, our results indicated that all proteins exhibit tighter binding (lower K_d ) as the proportion of anionic lipid increases. Interestingly, at high proportions of phosphatidylserine, the K_d values of all four proteins began to converge. We also found that although k_off values for all four proteins followed trends similar to those observed for the K_d values, the variation among the proteins was much lower, indicating that much of the variation came from the kinetic binding (k_on) of the proteins. These findings indicate that the combination of silicon photonic microring resonator arrays and nanodiscs enables rapid interrogation of biomolecular binding interactions at model cell membrane interfaces.

SUBMITTER: Muehl EM 

PROVIDER: S-EPMC5625054 | biostudies-literature | 2017 Sep

REPOSITORIES: biostudies-literature