Abstract

High-voltage activated calcium channels are modulated by a series of auxiliary proteins, including those of the α₂δ family. Until recently, only a single α₂δ subunit was known, but two further members, α₂δ-2 and -3, have since been identified. In this study, the structure of these two novel subunits has been characterized and binding of the antiepileptic drug gabapentin investigated. Using antibodies directed against the amino terminal portion of the proteins, the gross structure of the subunits could be analyzed by Western blotting. Similar to α₂δ-1, both α₂δ-2 and -3 subunits consist of two proteins—a larger α₂ and a smaller δ that can be separated by reduction. The subunits are also highly N-glycosylated with approximately 30 kDa of their mass consisting of oligosaccharides. α₂δ-1 was detected in all mouse tissues studied, whereas α₂δ-2 was found at high levels in brain and heart. The α₂δ-3 subunit was observed only in brain. α₂δ-1 and α₂δ-2, but not α₂δ-3, were found to bind gabapentin. TheK _d value of gabapentin binding to α₂δ-2 was 153 nM compared with the higher affinity binding to α₂δ-1 (K _d = 59 nM).