Sedimentation-Velocity AUC
Sedimentation-velocity experiments were performed in a Beckman Optima Xli analytical ultracentrifuge, using aluminum double-sector sapphire cells in an An-50 Ti rotor. The rotor speed was 50,000 rpm, and the temperature was maintained at 277 K. Prior to centrifugation, protein samples were dialyzed exhaustively against the buffer blank (50 mM Tris/HCl [pH 8.0], 300 mM NaCl, and 0.50 mM TCEP). Interference images were collected every 180 s during the sedimentation run. The data recorded from moving boundaries were analyzed by the program SEDFIT in terms of both discrete species and continuous distribution function of sedimentation coefficient (c(s)) and molar mass (c(M)), allowing determinations of sedimentation coefficient and shape-independent molecular weight (Brown and Schuck, 2006).