Crystal Structure of Human Tetraspanin CD81 Reveals a Conserved Intramembrane Binding Cavity

Abstract

Tetraspanins comprise a diverse family of four‐pass transmembrane proteins that play critical roles in cell activation, adhesion, migration and proliferation. CD81/Tspan28, the most widely studied member of the family, forms a complex with CD19, CD21 and CD225 to regulate B cell receptor function, and also acts as a receptor or co‐receptor for pathogens such as Hepatitis C virus and Plasmodium species. Despite the fundamental importance of tetraspanin proteins in a broad range of physiologic and pathophysiologic processes, structural information about a full‐length tetraspanin has remained elusive. Here we present the first crystal structure of a full‐length tetraspanin, human CD81. The transmembrane segments of CD81 pack as two largely separated pairs of helices, not as the four‐helix bundle predicted previously. The structure is capped by the large extracellular loop 2 (EC2) at the outer membrane leaflet, and the two pairs of helices converge at the inner leaflet to form a shape resembling an inverted teepee. The transmembrane region contains a large pocket suggestive of a ligand‐binding site, offering a potential mechanism for binding lipids or other molecules. In support of this idea, we observe electron density consistent with a bound cholesterol molecule in the pocket, and we detect specific binding of cholesterol to this site in vitro. The discovery of the hydrophobic pocket in the core of the transmembrane domain presents a novel therapeutic avenue for targeting CD81 and other tetraspanins in pathogenic infections or malignancies.Support or Funding InformationBrandon Zimmerman is supported by a CIHR postdoctoral fellowship. NIH grant NCI 5ROI CA092433 to SCB and DP5 OD021345 to ACK.