Abstract
The adhesion G protein–coupled receptor CD97 and its ligand complement decay-accelerating factor CD55 are important binding partners in the human immune system. Dysfunction in this binding has been linked to immune disorders such as multiple sclerosis and rheumatoid arthritis, as well as various cancers. Previous literatures have indicated that the CD97 includes 3 to 5 epidermal growth factor (EGF) domains at its N terminus and these EGF domains can bind to the N-terminal short consensus repeat (SCR) domains of CD55. However, the details of this interaction remain elusive, especially why the CD55 binds with the highest affinity to the shortest isoform of CD97 (EGF1,2,5). Herein, we designed a chimeric expression construct with the EGF1,2,5 domains of CD97 and the SCR1–4 domains of CD55 connected by a flexible linker and determined the complex structure by crystallography. Our data reveal that the two proteins adopt an overall antiparallel binding mode involving the SCR1–3 domains of CD55 and all three EGF domains of CD97. Mutagenesis data confirmed the importance of EGF5 in the interaction and explained the binding specificity between CD55 and CD97. The architecture of CD55–CD97 binding mode together with kinetics suggests a force-resisting shearing stretch geometry when forces applied to the C termini of both proteins in the circulating environment. The potential of the CD55–CD97 complex to withstand tensile force may provide a basis for the mechanosensing mechanism for activation of adhesion G protein–coupled receptors.
Highlights
Adhesion G protein–coupled receptors (GPCRs) are a subfamily of GPCRs that participate in a wide variety of functions from cell adhesion to immune defense and development, and, their dysfunction is linked to a myriad of negative health effects including inflammation, neurological disease, and cancer [1]
The CD55–CD97 pair has been indicated to play an important role in host defense and inflammation, as they can mediate cell adhesion and prevent the uncontrolled clustering of leukocytes in the blood stream [20, 32]
Despite a slow on-rate, CD97 is in close proximate distance to CD55 during cell adhesion and clustering; thereby, fast formation of a linkage between these two molecules is feasible as a result of high local concentration
Summary
The CD55 SCR1–4 domains and CD97 EGF1,2,5 domains were initially purified independently from HEK293 cells. EGF1 forms extensive hydrophobic interactions with SCR2 and SCR3 residues and buries a total surface area of 518 Å2. The preeminent interactions of CD97– CD55 come from the EGF2–SCR1/SCR2 interface in the middle region, which buries a total surface area of 1278 Å2 and contains six pairs of hydrogen bonds (Fig. 2B). At the bottom half of the interface, the EGF2–SCR1 interactions are secured by three pairs of side chain–main chain hydrogen bonds between EGF2 residues Asp-79, Asp-80, and Asp-81 and SCR1 residues Asp-77, Lys-71, and Lys-74, respectively. In the third interface of EGF5–SCR1 (Fig. 2C), Val-137 of EGF5 inserts into a joint hydrophobic pocket formed by SCR1 residues Val-60, Lys-76, Val-79, Ile-80, and Leu-82 and EGF2 residue Phe-79 These interfaces are consistent with previous literature, identifying the EGF1,2 and SCR1,2 domains as critical determinants for CD55–CD97 interactions. Our structure unveils additional interactions contributed by EGF5 of CD97 as well as EGF1-SCR2/SCR3 interface
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