Movement of small ensembles of motors bound to a Qdot, visualized by TIRF microscopy. (A) Kymographs of movement of multiple motors (either Myo51 or Myo51–Rng8/9) bound to a Qdot on either actin or actin-Tpm tracks. Note that Myo51–Rng8/9 shows no motion on actin-Tpm. (B) Histograms of run lengths of Myo51 on actin (n = 231) or actin-Tpm (n = 221), and Myo51–Rng8/9 on actin (n = 288). The red lines are exponential fits (y = Ae^−x/λ) to the data. The run length constants λ are indicated. The run length of Myo51 is higher on actin-Tpm compared with bare actin (P < 0.01, Mann–Whitney test). (C) Normalized run frequencies and run speed distributions. The mean run frequency of Myo51 on bare actin (number of events per µm actin per µM Myo51 per second) was normalized to one. Mean speed is indicated by the red lines. Mean ± SD values are indicated. Myo51 run speed is slower on actin-Tpm compared with bare actin (P < 0.001, Mann–Whitney test). On bare actin, the presence of Rng8/9 bound to Myo51 caused no statistical difference in run length, run frequency, or speed.