Biochemistry and Biophysics of Molecular Motors

Biochemistry and Biophysics of Molecular Motors



This series of papers is intended for an introductory biochemistry course for an intensive integrated science program with students who have strong backgrounds in mathematics, and physics. However, they could also be appropriate for introductory or intermediate biochemistry and biophysics courses, as well as other advanced biology courses that focus on proteins.

The papers focus on the biochemistry and biophysics of molecular motors and the elucidation of the detailed mechanisms that describe how these motors move on microtubules. These papers integrate many disciplines, including biochemistry, biophysics, cell biology, genetics, molecular biology, and statistics. Methods across these disciplines are used, and central concepts in biochemistry can be learned from these papers.

The papers have clear styles of writing despite the complexity of the experimental systems and data analyses. The questions, hypotheses, experimental designs, data analyses, and conclusions follow from one another and are typically well defined.

This series of papers also track the professional development of two first authors from a graduate student and a postdoctoral fellow to now faculty members with their independent research programs. The earlier papers are separate but related research projects from one large group, and the later papers are written by the first authors after they become independent principle investigators.


Applicable for Courses:

Biochemistry and Biophysics

Educational Level:

Variety of levels

Roadmap Objectives:

    • Article: Yildiz A, Tomishige M, Vale RD, Selvin PR (2004). Kinesin walks hand-over-hand. Science 303: 676-8.
    • Content area/major concepts: In this paper, the hypothesis is well laid out, the experiment and data are well described, and the conclusion is quite simple. It sets the stage for subsequent investigations that extend from a simple experiment to detailed mechanistic studies and from kinesin to dynein, a similar but much more complex protein machinery.

      gene expression, protein structure, protein structure determination, chemical and physical properties of macromolecules, modularity of proteins, molecular motors, conformational change, ATP hydrolysis (kinetics and thermodynamics), randomness and stochasticity, equilibrium, hypothesis and model testing
    • Methods or technology used to obtain data: molecular cloning, site-directed mutagenesis, protein expression, protein purification, affinity tags, chemical cross-linking, fluorescence probe, protein structure modeling, total internal reflection fluorescence (TIRF) microscopy, fluorescence imaging one-nanometer accuracy (FIONA), statistical and probabilistic modeling
    • How the CREATE strategy was used:
    • Biggest teaching challenge: Because of the seemingly simplicity of the paper, students might think that they understand the paper without realizing the underlying complexity of the concepts and methods involved.
    • Article: Yildiz A, Tomishige M, Gennerich A, Vale RD (2008). Intramolecular strain coordinates kinesin stepping behavior along microtubules. Cell 134: 1030-41.
    • Content area/major concepts: This second paper extends the basic experiment in the previous paper and examines how kinesin walks on microtubules and how conformational flexibility in kinesin can affect its movement. It ties protein
      conformation to function at a level that requires deep understanding of macromolecular properties while maintaining the familiarity of the experimental system.

      conformational flexibility, conformational strain, unstructured protein regions, antibody, and related material from previous paper
    • Methods or technology used to obtain data: enzymatic assay, centrifugation, spectroscopy, optical tweezer, antibody capture, and related methods from previous paper
    • How the CREATE strategy was used:
    • Biggest teaching challenge: The mathematics and physics involved might also be challenging for students who might not have extensive backgrounds in these areas.
    • Article: Reck-Peterson SL, Yildiz A, Carter AP, Gennerich A, Zhang N, Vale RD (2006). Single-molecule analysis of dynein processivity and stepping behavior. Cell 126: 335-48.
    • Content area/major concepts: This paper extends the investigation from kinesin to dynein and introduces a number of more complex experimental methods to study a protein machinery that is much more complex than kinesin. Enzyme mechanisms and enzyme kinetics can be introduced through this paper because of its use of TEV protease and HaloTag (a modified haloalkane dehalogenase).

      homologous recombination, protease, enzyme mechanism, enzyme kinetics, cofactor, protein-ligand interaction, orthogonal biochemical processes, and related material from previous papers
    • Methods or technology used to obtain data: yeast genetics, inducible gene expression system, electrophoresis, Western blot, HaloTag, biotin-streptavidin capture, gradient centrifugation, and related methods from previous papers
    • How the CREATE strategy was used:
    • Biggest teaching challenge: This paper can be a big leap from the previous two, because dynein is a much more complex protein, and many more complicated experiments are involved.
    • Article: Derr ND, Goodman BS, Jungmann R, Leschziner AE, Shih WM, Reck-Peterson SL (2012). Tug-of-war in motor protein ensembles revealed with a programmable DNA origami scaffold. Science 338: 662-5.
    • Content area/major concepts: This paper completes the loop by tying kinesin and dynein together in one experimental system to analyze how the interplay between the two proteins can affect cargo transport. The experimental system also emphasizes the complex nature of biological phenomena.

      nucleic acids, DNA complementarity, reaction kinetics, complex system (multiple macromolecules acting at the same time), and related material from previous papers
    • Methods or technology used to obtain data: electrophoretic mobility shift assay, electron microscopy (protein structure), photocleavable probe, and related methods from previous papers
    • How the CREATE strategy was used:
    • Biggest teaching challenge: The experimental system can be challenging to understand. The use of DNA as an experimental tool can also be confusing, because it is used in a context different from its usual function.

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