(a) Top view of the LBD layer of the CA structure23. Subunits A (green) and C (blue) are covalently linked by a disulfide bond formed between the introduced Cys665 on subunits A and C. The lower panel cartoon shows the localization of the position A665C. (b) Patch clamp experiments showing test pulses for cysteine mutations on positions 664-666. CTZ (100 μM) was present throughout the whole experiment. The recovery of the current in 10mM glutamate and DTT following trapping in the presence of CuPhen (10 μM) with 5-fluorowillardiine (FW) (right panel) or Kainate (KA) (left panel) was recorded. Arrows indicate a reduction of the current after trapping, which was observed for I664C, A665C and V666C but not for wild type (WT, bottom panel). Open circles indicate double exponential fits to the recovery after trapping. The time constants are summarized in Fig. 5. The difference between the active fraction after trapping in the presence of FW and KA for I664C, A665C and V666C compared with WT was significant for FW (P<0.005) and KA (P<0.05).