Figure 7
(A,B) Sedimentation velocity experiments for detection of interaction between phLDH and byGAPDH. The sedimentation profiles for a mixture of phLDH and byGAPDH were measured by following scans at 280 nm. The two-enzyme mixture was prepared using 6 μM solution in 50 mM Tris/HCl, pH = 7.2, 1 mM EDTA, 0.5 mM DTT. In both panels measured sedimentation profiles are shown in black, and best-fit profiles are shown in light gray lines. (A) the panel shows overlap between sedimentation profiles and the best-fit profiles assuming one component with sedimentation constant s020,w = 7.6 ± 0.4 S, Mr = 143,6 ± 0.9 kDa. (B) the panel shows measured sedimentation profiles for the two proteins mixture just as panel A, except that the gray lines show Claverie simulation profiles for a single component with 10% self-association. The observed differences between measured sedimentation profiles and the calculated profiles indicate that there is no detectable interaction between phLDH and byGAPDH with the KD constant lower than 48.6 μM.
