Interactions between Thrombomodulin and the Complement System Studied by Surface Plasmon Resonance and Deuterium Exchange Mass Spectrometry

Abstract

Complement component 3 (C3) is at the junction of three different complement activation pathways (classical, lectin, and alternative). Activation to C3b is a key step in the innate immune response that allows for the formation of important multi-protein complexes that ultimately participate in pathogen clearance. When misregulated, however, complement can lead to inflammatory disease and autoimmune disorders. Several regulatory proteins for C3b are known, but the molecular details of interactions between these proteins have not yet been elucidated. Thrombomodulin (TM), and specifically its N-terminal lectin-like domain (TMD1), has been identified as a possible regulator of complement through interactions with C3 or C3b, and the known regulator CFH may also be required. We have used surface plasmon resonance (SPR) and hydrogen/deuterium exchange mass spectrometry (HDXMS) to study the interaction of TMD1 with C3 and C3b. Using SPR, we see that C3 binds to a surface coated with TMD1, and full kinetic studies are underway. Using HDXMS, we see that TMD1 interacts with C3b, and there is a lesser interaction with C3. TMD1 tends to make C3b more accessible to deuterium exchange, while C3 tends to be less accessible to deuterium exchange in the presence of TMD1. This difference suggests a possible role for TMD1 in regulating a key step along the complement pathway. We will next investigate the role of CFH in these interactions.