FIGURE 4.
A, wild-type β2m NMR DOSY spectra as a function of α-crystallin concentration and time. A limited window of the aliphatic region is shown to highlight the small but significant alteration in diffusion coefficient. The contour traces of five different DOSY experiments are superimposed at α-crystallin/β2m molar ratios of 0:1 (black), 1:4.9 × 102 (green), 1:95 (blue), 1:35 (pink), and 1:35 after 230 days from the last α-crystallin addition (red). The substantial overlap of the traces at the two lowest and highest α-crystallin concentrations reflects the increase in translational diffusion coefficient value, DT, of β2m below some 100-fold molar excess with respect to α-crystallin. The effect persists with solution aging. B, corresponding DT values of β2m as a function of α-crystallin concentration. The plotted data are the center of gravity measured at 1.6 ppm in the DOSY traces in A, with errors representing the maximum excursions above and below the determined value in the same traces. The red point refers to the measurement repeated 230 days after the first determination on the freshly prepared solution of 1:35 α-crystallin/β2m.