Kinesin Protocols

Buch | Softcover

258 Seiten

2010 | Softcover reprint of hardcover 1st ed. 2001
Humana Press Inc. (Verlag)
978-1-61737-178-3 (ISBN)

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By the end of the 1980s only two microtubule-dependent motors, the plus end-directed kinesin and the minus end-directed cytoplasmic dynein, had been identified. At the time, these two motors seemed almost sufficient to explain directional motility events on polar microtubule tracks in the cell. No- theless, shortly after, the tip of the iceberg began to emerge with the identi- cation of proteins containing in their sequences a domain found in kinesin. This domain, called the “motor domain,” conferred on these proteins the essential property of moving on microtubules, using the energy derived from ATP hydro- sis. Since then, the identification of new proteins belonging to the kinesin superfamily of microtubule-dependent motors has gone at such a pace that nowadays more than 200 entries with motor domain sequences are deposited in the database. Kinesin family members are found in all eukaryotic org- isms tested. They present a wide range of domain organizations with a motor domain located at different positions in the molecule. Their motility prop- ties are also variable in directionality, velocity, and such other characteristics as bundling activity and processivity. Finally, and most important, they p- ticipate in a multitude of cellular functions. Our understanding of many cel- lar events, such as mitotic spindle assembly and neuronal transport, to cite only two, has progressed substantially in the last few years thanks to the id- tification of these motors.

Purification of Kinesin from the Brain.- RT-PCR for the Identification of Developmentally Regulated Novel Members of the Kinesin-like Superfamily.- Expression Cloning with Pan Kinesin Antibodies.- Expression of Kinesin in Escherichia coli.- Plasmids for Expression of Chimeric and Truncated Kinesin Proteins.- Preparation of Recombinant Kinesin Superfamily Proteins Using the Baculovirus System.- Assays for Kinesin Microtubule-Stimulated AT Pase Activity.- An Improved Microscope for Bead and Surface-Based Motility Assays.- Use of Photonic Force Microscopy to Study Single-Motor-Molecule Mechanics.- Assays for Microtubule-Destabilizing Kinesins.- Green Fluorescent Protein as a Tag for Molecular Motor Proteins.- In Vitro Reconstitution of Endosome Motility Along Microtubules.- Approaches to Study Interactions Between Kinesin Motors and Membranes.- Microinjection Methods for Analyzing the Functions of Kinesins in Early Embryos.- The Use of Dominant Negative Mutants to Study the Function of Mitotic Motors in the In Vitro Spindle Assembly Assay in Xenopus Egg Extracts.- A Dominant Negative Approach for Functional Studies of the Kinesin II Complex.- Identification of Kinesin-Associated Proteins.- Assaying Spatial Organization of Microtubules by Kinesin Motors.- Crystallization of Kinesin.- Structural Analysis of the Microtubule–Kinesin Complex by Cryo-Electron Microscopy.

"This is a concise but comprehensive book that would be a valuable reference for any researcher who wishes to study this important cytoplasmic motor protein. It would be a valuable book to have in the laboratory as well as a useful reference book in a life sciences library. "-Doody's Health Sciences Book Review Journal

Erscheint lt. Verlag	10.11.2010
Reihe/Serie	Methods in Molecular Biology ; 164
Zusatzinfo	XI, 258 p.
Verlagsort	Totowa, NJ
Sprache	englisch
Maße	152 x 229 mm
Themenwelt	Naturwissenschaften ► Biologie ► Mikrobiologie / Immunologie
Themenwelt	Naturwissenschaften ► Biologie ► Zellbiologie
ISBN-10	1-61737-178-5 / 1617371785
ISBN-13	978-1-61737-178-3 / 9781617371783
Zustand	Neuware