Kinesin Protocols

1. Front Matter

   Pages i-xi

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2. Purification of Kinesin from the Brain

   • Sergei A. Kuznetsov, Vladimir I. Gelfand

   Pages 1-7

3. RT-PCR for the Identification of Developmentally Regulated Novel Members of the Kinesin-like Superfamily

   • Niovi Santama

   Pages 9-20

4. Expression Cloning with Pan Kinesin Antibodies

   • Laura M. Ginkel, Linda Wordeman

   Pages 21-41

5. Expression of Kinesin in Escherichia coli

   • Maryanne F. Stock, David D. Hackney

   Pages 43-48

6. Plasmids for Expression of Chimeric and Truncated Kinesin Proteins

   • Kimberly W. Waligora, Sharyn A. Endow

   Pages 49-55

7. Preparation of Recombinant Kinesin Superfamily Proteins Using the Baculovirus System

   • Nobutaka Hirokawa, Yasuko Noda

   Pages 57-63

8. Assays for Kinesin Microtubule-Stimulated AT Pase Activity

   • David D. Hackney, Wei Jiang

   Pages 65-71

9. An Improved Microscope for Bead and Surface-Based Motility Assays

   • Nick Carter, Rob Cross

   Pages 73-89

10. Use of Photonic Force Microscopy to Study Single-Motor-Molecule Mechanics

    • Sylvia Jeney, Ernst-Ludwig Florin, J. K. Heinrich Hörber

    Pages 91-108

11. Assays for Microtubule-Destabilizing Kinesins

    • Arshad Desai, Claire E. Walczak

    Pages 109-121

12. Green Fluorescent Protein as a Tag for Molecular Motor Proteins

    • Sharyn A. Endow

    Pages 123-131

13. In Vitro Reconstitution of Endosome Motility Along Microtubules

    • Erik Nielsen, Fedor Severin, Anthony A. Hyman, Marino Zerial

    Pages 133-146

14. Approaches to Study Interactions Between Kinesin Motors and Membranes

    • Gerardo Morfini, Ming-Ying Tsai, Györgyi Szebenyi, Scott T. Brady

    Pages 147-162

15. Microinjection Methods for Analyzing the Functions of Kinesins in Early Embryos

    • Robert L. Morris, Heather M. Brown, Brent D. Wright, David J. Sharp, William Sullivan, Jonathan M. Scholey

    Pages 163-172

16. The Use of Dominant Negative Mutants to Study the Function of Mitotic Motors in the In Vitro Spindle Assembly Assay in Xenopus Egg Extracts

    • Haralabia Boleti, Eric Karsenti, Isabelle Vernos

    Pages 173-189

17. A Dominant Negative Approach for Functional Studies of the Kinesin II Complex

    • Vladimir I. Gelfand, Nathalie Le Bot, M. Carolina Tuma, Isabelle Vernos

    Pages 191-204

18. Identification of Kinesin-Associated Proteins

    • Lisa C. Lindesmith, Janardan Kumar, Michael P. Sheetz

    Pages 205-212

19. Assaying Spatial Organization of Microtubules by Kinesin Motors

    • François Nédélec, Thomas Surrey

    Pages 213-222

20. Crystallization of Kinesin

    • Manfred Thormählen, Jens Müller, Eckhard Mandelkow

    Pages 223-233

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.

"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

• European Molecular Biology Laboratory, Heidelberg, Germany

  Isabelle Vernos

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