Purification of axonemal dyneins and dynein-associated components from Chlamydomonas.
Axonemal dyneins are responsible for generating the force required to power ciliary and flagellar motility. These highly complex enzymes form the inner and outer arms associated with the outer doublet microtubules. They are built around one or more ~520kD heavy chains that exhibit motor activity and also include additional components that are required for assembly within the axonemal superstructure and/or regulation of motor function in response to a broad range of signaling inputs. The dyneins from flagella of Chlamydomonas have been extensively studied as this organism is amenable to genetic, biochemical, and physiological approaches. In this chapter, I describe methods that have been devised by a number of laboratories to extract and purify individual dyneins from Chlamydomonas. When combined with the wide range of available mutants, these methods allow for the analysis of dyneins lacking individual components or motor units.
King
Stephen M
SM
Department of Molecular, Microbial and Structural Biology, University of Connecticut Health Center, Farmington, 06030-3305, USA.
D020418:Algal Proteins / Q000302:isolation & purification* / Q000378:metabolism; D056444:Axonemal Dyneins / Q000737:chemistry / Q000302:isolation & purification*; D001671:Biochemistry / Q000379:methods*; D002457:Cell Extracts; D002499:Centrifugation, Density Gradient; D005591:Chemical Fractionation; D002696:Chlamydomonas / Q000378:metabolism*; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D005407:Flagella / Q000378:metabolism; D047468:Immunoprecipitation; D009154:Mutation / Q000235:genetics; D013347:Subcellular Fractions / Q000378:metabolism
D020418:Algal Proteins; D002457:Cell Extracts; D056444:Axonemal Dyneins