Crystal data processing and structure determination

Harvested crystals were soaked in 25% glycerol before being carefully fished with cryoloops and flash-frozen in liquid nitrogen for subsequent data collection at the National Synchrotron Radiation Research Center, Taiwan. Indexing, scaling, and merging of datasets were carried out in HKL2000 (55). The crystal structure twinfilin/CP/actin complex was solved at a resolution of 3.2 Å by molecular replacement using Phaser (56), sequentially searching for one copy of mouse twinfilin-1 D2 (3DAW) (19) and one copy of chicken CP heterodimer (3AA7) (35) followed by a second copy of mouse twinfilin-1 D2 (3DAW) (19). The resultant model was rebuilt by hand, and the structure was subjected to several rounds of refinement using Phenix (56) and further manual rebuilding in COOT (57). The crystal data collection and structure refinement statistics are presented in table S1.