Purification and characterization of Chlamydomonas flagellar dyneins
Department of Cell Biology
Adenosine Triphosphatases; Ca(2+) Mg(2+)-ATPase; Centrifugation, Density Gradient; Chlamydomonas; Chromatography; Durapatite; Dynein ATPase; Flagella; Hydroxyapatites; Indicators and Reagents; Molecular Weight
Algae | Amino Acids, Peptides, and Proteins | Cell Biology | Enzymes and Coenzymes
Dyneins are ATPases which transduce the chemical energy of ATP into the mechanical force required for ciliary and flageUar beating, i These enzymes are located in the inner and outer arms that project from the outer doublet microtubules of cilia and flagella. Dyneins have been ex- tracted and partially or completely purified from a number of different sources, including cilia of the protozoans Tetrahymena 2 and Parame- cium,3 cilia of lamellibranch gills (Aequipecten 4 and UnioS), sperm flagella of starfish (Asterias6), trout (Salmo7), bull (Bos8.9), and a variety of sea urchins (e.g., Colobocentrotus,10 Hemicentrotus,11 Strongylocentrotus, 12 and Tripneustes~3), and flagella of the green alga Chlamydomonas. 14~5 Chlamydomonas has several advantages for the purification and study of dynein. First, large quantities of Chlamydomonas are easily grown synchronously 16,17 in axenic defined medi#8; second, the flagella are read- ily detached and separated from the cell bodies19.2°; third, highly purified dyneins can be obtained with a minimum of proteolysis15; fourth, the synthesis of dynein polypeptides and their assembly into newly formed arms can be studied during flagellar regeneration21; fifth, the dynein poly- peptides can be readily labeled in vivo with 3H,22 32p,23,24 or 35S14,25,26; and finally, the isolation and biochemical characterization of axonemes pre- pared from mutant Chlamydomonas defective in their inner and/or outer arms 27,2~ permit elegant and detailed analysis of the composition, assem- bly, and function of their dynein components. As a result, a wealth of information is now available on the structural, biochemical, and physical properties of Chlamydomonas dyneins.
DOI of Published Version
Methods Enzymol. 1986;134:291-306.
Methods in enzymology
King SM, Otter T, Witman GB. (1986). Purification and characterization of Chlamydomonas flagellar dyneins. Cell and Developmental Biology Publications. https://doi.org/10.1016/0076-6879(86)34097-7. Retrieved from https://escholarship.umassmed.edu/cellbiology_pp/63