Figure 2

MALLS-SEC analyses of MCR1 Fractions A, B, and C. (A) Representative elution profiles of each fraction from SEC (20 ml Superdex 200 gel filtration column) and monitored by refractometry. Only the peaks retained during chromatographic separation are shown. Samples were incubated at 10°C for 30 min prior to analysis. (B) Molar mass distribution plots for each fraction. The elution profiles for each fraction in panel A are shown at full scale (including sample eluting in the void volume). The molar masses for sample eluting in the retained peaks are shown as filled circles. The molar mass is shown on the y-axis in log scale. Top panel: The elution profile of Fraction A consisted of two retained peaks with experimental masses of 520 kDa and 280 kDa, consistent with tetrameric and dimeric MCR1, respectively. (Due to incomplete resolution of peaks, the experimental masses tend to be ~10–20% larger than the predicted masses of the MCR1 oligomeric forms.) Middle panel: The retained peaks in elution profile of Fraction B included a broad peak, which was polydisperse in molecular mass, likely consisting of tetrameric and higher order oligomers of MCR1. A second retained peak was present, with experimental mass of 290 kDa, consistent with dimeric MCR1. Bottom panel: The elution profile of Fraction C consisted of a retained peak with experimental mass of 990 kDa, consistent with octameric MCR1.