Structure and Functional Expression of an ω-Conotoxin-Sensitive Human N-Type Calcium Channel

Abstract

N-type calcium channels are ω-conotoxin (ω-CgTx)-sensitive, voltage-dependent ion channels involved in the control of neurotransmitter release from neurons. Multiple subtypes of voltage-dependent calcium channel complexes exist, and it is the α 1 subunit of the complex that forms the pore through which calcium enters the cell. The primary structures of human neuronal calcium channel α 1B subunits were deduced by the characterization of overlapping complementary DNAs. Two forms (α 1B-1 and α 1B-2 ) were identified in human neuroblastoma (IMR32) cells and in the central nervous system, but not in skeletal muscle or aorta tissues. The α 1B-1 subunit directs the recombinant expression of N-type calcium channel activity when it is transiently co-expressed with human neuronal β 2 and α 2b subunits in mammalian HEK293 cells. The recombinant channel was irreversibly blocked by ω-CgTx but was insensitive to dihydropyridines. The α 1B-1 α 2b β 2 -transfected cells displayed a single class of saturable, high-affinity (dissociation constant = 55 pM) ω-CgTx binding sites. Co-expression of the β 2 subunit was necessary for N-type channel activity, whereas the α 2b subunit appeared to modulate the expression of the channel. The heterogeneity of α 1b subunits, along with the heterogeneity of α 2 and β subunits, is consistent with multiple, biophysically distinct N-type calcium channels.