Ca2+ permeability of the (alpha4)3(beta2)2 stoichiometry greatly exceeds that of (alpha4)2(beta2)3 human acetylcholine receptors.
Human alpha4beta2 nicotinic acetylcholine receptors (AChRs) expressed in Xenopus laevis oocytes or transfected cell lines are present as a mixture of two stoichiometries, (alpha4)2(beta2)3 and (alpha4)3(beta2)2, which differ depending on whether a beta2 or alpha4 subunit occupies the accessory subunit position corresponding to beta1 subunits of muscle AChRs. Pure populations of each stoichiometry can be expressed in oocytes by combining a linked pair of alpha4 and beta2 with free beta2 to produce the (alpha4)2(beta2)3 stoichiometry or with free alpha4 to produce the (alpha4)3(beta2)2 stoichiometry. We show that the (alpha4)3(beta2)2 stoichiometry and the (alpha4)2(beta2)2beta3 and (alpha4)2(beta2)2alpha5 subtypes in which beta3 or alpha5occupy the accessory positions have much higher permeability to Ca2+ than does (alpha4)2(beta2)3 and suggest that this could be physiologically significant in triggering signaling cascades if this stoichiometry or these subtypes were found in vivo. We show that Ca2+ permeability is determined by charged amino acids at the extracellular end of the M2 transmembrane domain, which could form a ring of amino acids at the outer end of the cation channel. Alpha4, alpha5, and beta3 subunits all have a homologous glutamate in M2 that contributes to high Ca2+ permeability, whereas beta2 has a lysine at this position. Subunit combinations or single amino acids changes at this ring that have all negative charges or a mixture of positive and negative charged amino acids are permeable to Ca2+. All positive charges in the ring prevent Ca2+ permeability. Increasing the proportion of negative charges is associated with increasing permeability to Ca2+.
Tapia
L
L
Departmento de Farmacología y Terapéutica, Facultad de Medicina, Universidad Autónoma de Madrid, Spain.
Kuryatov
A
A
Lindstrom
J
J
D000109:Acetylcholine / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D005260:Female; D006801:Humans; D008969:Molecular Sequence Data; D010539:Permeability; D011978:Receptors, Nicotinic / Q000737:chemistry* / Q000145:classification / Q000502:physiology; D013329:Structure-Activity Relationship; D014982:Xenopus laevis
D011978:Receptors, Nicotinic; C403531:nicotinic receptor alpha4beta2; D000109:Acetylcholine; D002118:Calcium