• Login as Editor
    View Item 
    •   Yeshiva Academic Institutional Repository
    • Albert Einstein College of Medicine (AECOM)
    • Albert Einstein College of Medicine: Doctoral Dissertations
    • View Item
    •   Yeshiva Academic Institutional Repository
    • Albert Einstein College of Medicine (AECOM)
    • Albert Einstein College of Medicine: Doctoral Dissertations
    • View Item
    JavaScript is disabled for your browser. Some features of this site may not work without it.

    ALLOAFFINITY PURIFICATION OF DYNEIN (CILIA, MT-ASSOCIATED ATPASE)

    Thumbnail

    Date
    1984
    Author
    NASR, ANTHONY
    Metadata
    Show full item record
    Share
    Abstract
    The bending movements of eukaryotic flagella and cilia have been shown to originate in a sliding motion between adjacent microtubule doublets. This sliding force can be attributed to the ciliary ATPase, dynein. Dynein can be extracted from the axonemes of Tetrahymena thermophila by low ionic strength dialysis. The soluble dynein will rebind to microtubules in an ATP sensitive way if the appropriate ionic conditions are reestablished. This phenomenon forms the basis of a rapid and highly specific purification procedure based on the ATP-regulated binding and release cycle.;Tetrahymena ciliary doublet microtubules, extracted free of dynein and other axonemal components, can be retained on the surface of a GVWP filter (Millipore). Buffer (+OR-)ATP passed through the microtubule-loaded filter yields neither microtubules nor ATPase in the filtrate. Crude dynein-containing extract (in the absence of microtubules) is able to pass through the filter unaffected, but when the dynein is allowed to bind to the microtubules before application to the filter, the dynein is retained. If buffer + ATP is now passed through the filter, the dynein is released into the filtrate. Since the technique demands the dynein to recognise two ligands, the procedure has been named alloaffinity chromatography.;Alloaffinity purified dynein from Tetrahymena has been extensively characterised. It is a monodisperse oligomer consisting of about 11 polypeptides, ATPase activity highly specific for ATP and, at physiological pH, the purified dynein displays non-linear saturation kinetics suggestive of negative cooperativity. The activity is stimulated 40% by the presence of microtubules. An adenylate kinase activity copurifies with the dynein in significant and reproducible amounts. Alloaffinity purified dynein is capable of rebinding to microtubules with a morphology appropriate for the in situ dynein arm. Thus, alloaffinity chromatography yields an authentic dynein molecule, suitable for further study. Alloaffinity chromatography has been applied to a crude flagellar extract of Chlamydomonas. Using Tetrahymena microtubules, the alloaffinity procedure yields a group of polypeptides which correspond well to the reported subunit composition of Chlamydomonas outer arm dynein. Alloaffinity chromatography is therefore potentially applicable to the study of axonemal dyneins from many species. In addition, the technique is a promising tool to search for dynein like molecules in non ciliated cells.
    Permanent Link(s)
    https://ezproxy.yu.edu/login?url=http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqm&rft_dat=xri:pqdiss:8417635
    https://hdl.handle.net/20.500.12202/2953
    Citation
    Source: Dissertation Abstracts International, Volume: 45-05, Section: B, page: 1454.
    *This is constructed from limited available data and may be imprecise.
    Collections
    • Albert Einstein College of Medicine: Doctoral Dissertations [1674]

    Yeshiva University Libraries copyright © 2021  DuraSpace
    YAIR Self-Deposit | YAIR User's Guide | Take Down Policy | Contact Us
    Yeshiva University
     

     

    Browse

    AllCommunities & CollectionsBy Issue DateAuthorsTitlesSubjectsThis CollectionBy Issue DateAuthorsTitlesSubjects

    My Account

    Login as Editor

    Statistics

    View Usage Statistics

    Yeshiva University Libraries copyright © 2021  DuraSpace
    YAIR Self-Deposit | YAIR User's Guide | Take Down Policy | Contact Us
    Yeshiva University