rat 42 and mouse ( 1)21 ion channels were expressed in xenopus laevis oocytes . for 42 receptors , subunit stoichiometry was controlled by varying the 4:2 subunit ratio and verified by voltage jump experiments . dose - response measurements for these channels were performed with a holding potential of 60 mv . unnatural amino acids and -hydroxy acids were prepared , coupled to dca and ligated to 74-mer thg73 as described previously15 . single - channel recording was performed in the cell - attached configuration with a pipette potential of + 100 mv as described previously27 . popen values were calculated from event - detected data using clampfit 9.2 single - channel search . full methods and any associated references are available in the online version of the paper at www.nature.com/nature . rat 42 and mouse ( 1)21 ion channels were expressed in xenopus laevis oocytes . for 42 receptors , subunit stoichiometry was controlled by varying the 4:2 subunit ratio and verified by voltage jump experiments . dose - response measurements for these channels were performed with a holding potential of 60 mv . unnatural amino acids and -hydroxy acids were prepared , coupled to dca and ligated to 74-mer thg73 as described previously15 . single - channel recording was performed in the cell - attached configuration with a pipette potential of + 100 mv as described previously27 . popen values were calculated from event - detected data using clampfit 9.2 single - channel search . full methods and any associated references are available in the online version of the paper at www.nature.com/nature .