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Isolation and characterization of plant myosin from pollen tubes of lily

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Summary

A plant myosin was isolated from pollen tubes of lily,Lilium longiflorum. Pollen tubes were homogenized in low ionic strength solution containing casein, and myosin from this crude extract was purified by co-precipitation with F-actin prepared from chicken breast muscle, followed by hydroxylapatite column and gel filtration column chromatography. Upon SDS-PAGE on 6% polyacrylamide gel, only 170 kDa polypeptide was detected in the purified myosin fraction. Furthermore, with immunoblotting using antiserum raised against 170 kDa polypeptide, only the 170 kDa component crossreacted in the crude sample of pollen tube proteins. This antiserum did not crossreact with the heavy chain of skeletal muscle myosin. The ATPase activity of pollen tube myosin was stimulated up to 60-fold by F-actin prepared from chicken breast muscle. The translocation velocity of rhodamine-phalloidin-labeled F-actin on a glass surface covered with pollen tube myosin ranged from 6.0 to 9.8 μm/s with an average of 7.7 μm/s. This velocity was similar to or a little faster than that of the cytoplasmic streaming that occurred in pollen tubes. These results suggested that myosin composed of a 170 kDa heavy chain produces the motive force for cytoplasmic streaming in pollen tube of lily.

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Abbreviations

ATP:

adenosine-5′-triphosphate

DTT:

dithiothreitol

EGTA:

ethyleneglycol-bis-(β-aminoethylether)N,N,N′,N′-tetraacetic acid

PAGE:

polyacrylamide gel electrophoresis

PIPES:

piperazin-N,N′-bis-(2-ethanesulfonic acid)

PMSF:

phenylmethylsulfonyl fluoride

SDS:

sodium dodecylsulfate

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Authors and Affiliations

  1. Department of Life Science, Faculty of Science, Himeji Institute of Technology, Harima Science Park City, 678-12, Hyogo, Japan

    E. Yokota & T. Shimmen

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  1. E. Yokota
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  2. T. Shimmen
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Yokota, E., Shimmen, T. Isolation and characterization of plant myosin from pollen tubes of lily. Protoplasma 177, 153–162 (1994). https://doi.org/10.1007/BF01378989

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  • DOI: https://doi.org/10.1007/BF01378989

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