Chemical crosslinking of mini-spectrin tetramers and dimers. (A) SDS-PAGE gel showing the mini-spectrin samples before and after crosslinking with EDC/NHS. The 60-minute time point is shown for each condition (complete time courses are shown in supplemental Figure 3). (B-D) Asterisk indicates the location of the L260P mutation, the dotted circle represents a distance of 50 Å from the N terminus end of the α0 helix that defines the maximum extension of the 20 residues in the disordered “tail” preceding the α0 helix; blue lines indicate approximate locations of crosslinks observed at 0°C and room temperature (RT); blue dots indicate sites to which the α-N terminus was observed to be crosslinked at 0°C and RT. The red lines represent crosslinks only observed at RT. (B) Crosslinks indicative of an open dimer conformation. (C) Crosslinks indicative of a closed dimer conformation. Circles are common to WT and L260P and + are specific to L260P. The red + is only observed at RT. (D) Crosslinks specific to the L260P dimer.