Discovery of ultrafast myosin, its amino acid sequence, and structural features.

Abstract

Cytoplasmic streaming with extremely high velocity (∼70 μm s^-1) occurs in cells of the characean algae (<i>Chara</i>). Because cytoplasmic streaming is caused by myosin XI, it has been suggested that a myosin XI with a velocity of 70 μm s^-1, the fastest myosin measured so far, exists in <i>Chara</i> cells. However, the velocity of the previously cloned <i>Chara corallina</i> myosin XI (<i>Cc</i>XI) was about 20 μm s^-1, one-third of the cytoplasmic streaming velocity in <i>Chara</i> Recently, the genome sequence of <i>Chara braunii</i> has been published, revealing that this alga has four myosin XI genes. We cloned these four myosin XI (<i>Cb</i>XI-1, 2, 3, and 4) and measured their velocities. While the velocities of <i>Cb</i>XI-3 and <i>Cb</i>XI-4 motor domains (MDs) were similar to that of <i>Cc</i>XI MD, the velocities of <i>Cb</i>XI-1 and <i>Cb</i>XI-2 MDs were 3.2 times and 2.8 times faster than that of <i>Cc</i>XI MD, respectively. The velocity of chimeric <i>Cb</i>XI-1, a functional, full-length <i>Cb</i>XI-1 construct, was 60 μm s^-1 These results suggest that <i>Cb</i>XI-1 and <i>Cb</i>XI-2 would be the main contributors to cytoplasmic streaming in <i>Chara</i> cells and show that these myosins are ultrafast myosins with a velocity 10 times faster than fast skeletal muscle myosins in animals. We also report an atomic structure (2.8-Å resolution) of myosin XI using X-ray crystallography. Based on this crystal structure and the recently published cryo-electron microscopy structure of acto-myosin XI at low resolution (4.3-Å), it appears that the actin-binding region contributes to the fast movement of <i>Chara</i> myosin XI. Mutation experiments of actin-binding surface loops support this hypothesis.