Figure 4.

Photolabeling of α₁β₃ GABA_A receptors by KK152 (the enantiomer of KK123). (A) The structure of KK152. (B) Extracted ion chromatograms from mass spectrometric analysis illustrating the labeling of α₁-TM4 peptide in α₁β₃ GABA_A receptors by KK152. The y-axis shows the intensity of the unlabeled TM4 peptide (top panel) and the KK152-labeled TM4 peptide (bottom panel). The x-axis shows the chromatographic retention time of the unlabeled TM4 peptide (blue, 105.9 min) and the KK152-labeled TM4 peptide (red, 108.5 min) illustrating that the increased hydrophobicity of the labeled peptide lengthens retention time on reversed phase chromatography. (C) HCD fragmentation spectrum of the α₁ subunit TM4 tryptic peptide photolabeled by 30 μM KK152. Red and black indicate fragment ions that do or do not contain KK152, respectively. The peptide sequence shows that the y-ion series of fragment ions contains diagnostic peptides indicating labeling of residue α₁ (Y415). (D) Structure of the α₁β₃ GABA_A receptor showing the location of Y415, the residue labeled by KK152 at the exoplasmic end of α₁ TM4. The α₁ subunit is shown in grey and the β₃ subunit in turquoise.