Fig 4. Effect of compound (1) or (2) on the oligomeric state of BcGR.

BcGR (33 μM) was incubated with 100 μM of compound (1) (Panel A) or (2) (Panel B) in the absence (red line) or in the presence of 10 mM D-Glu (blue line), and loaded on a Superdex 75 5/150 GL column, as in Materials and Methods. The peak corresponding to dimeric species (~70 kDa) is indicated with an arrow in both chromatograms. Integration of the peaks revealed a 2:1 monomer-dimer ratio for compound (1) (peak areas 1.6 x 10⁴ mV² and 4.4 x 10³ mV², for the monomer and dimer area, respectively), and 3:1 for compound (2) (peak areas 1.0 x 10⁴ mV² and 4.2 x 10³ mV², respectively). The additional 280 nm absorbance peak, corresponding to the column void volume, observed in the elution profile of BcGR incubated with compound (2) is presumably related only to the compound. This peak is not present in the fluorescence traces (S7 Fig), and no protein could be detected in corresponding fractions when loaded on SDS-PAGE (S8 Fig). Therefore, the content of this peak was not considered in further analyses.