Reading & Problems: LNC p. 179-183; Problems in sample study problems and practice final exam
I. Cytoskeletal fibers, tubulin/microtubule example
A. Microtubules are tubular polymers of alpha/beta tubulin dimers.
Microtubules have polarity, the end with exposed beta subunits is the +end and the end with the exposed alpha subunits is the -end (minus end).
B. Microtubules are dynamic, they can grow, or shrink. The rate of growth is the rate of tubulin dimer addition minus the rate of tubulin dimer loss. This is kon[tubulin]-koff.
C. The rate of growth can be different at the two ends so that microtubules can simultaneously grow at one end and shrink at the other in the process of "treadmilling".
D. Growth is regulated by binding and hydrolysis of GTP. GTP tubulin has higher affinity for tubule growth than does GDP tubulin. Tubulin can only hydrolyze GTP to GDP when it is within a microtubule. This can explain treadmilling. Here is an animation of microtubule assembly. More details on why the two ends behave differently
E. Tubulin adopts multiple different conformations depending on whether GTP or GDP is bound and whether it is bound to other tubulin dimers. Together these changes can explain microtubule dynamics including the reformation of microtubule arrays to move chromosomes during cell division.
- Kinesins move from - to + on microtubules. Some carry cargo from interior to exterior of cells. Others slide microtubules along each other and are involved in things like movement of chromosomes.
- Dyneins move from the + to the - ends of microtubules and carry cargo from the exterior to the interior of cells and are also involved in flagellar/cilliar motion.
- Myosins move on actin filaments and are involved in movement of cells and in muscle contraction.
B. How do they work? They use the energy of ATP hydrolysis to directionally drive conformational changes that allow them to "walk" along microtubules or actin filaments. The kinesin example is described below.
- Kinesins are homodimers with a coiled coil tether cargo-carrying domain and two "head" motor domains. The head domains are attached to the coiled coil by flexible "neck linker" domains.
- When not bound to microtubules (MTs) the head domains have ADP bound. This puts them in a conformation that has a high affinity for MT binding.
- When a head domain binds a MT, its conformation changes allowing release of ADP and binding of ATP.
- The binding of ATP causes a conformational change in the head domain that exposes a surface on the head that the neck region will "zipper" on to.
- This throws the other head forward where it can bind to the MT.
- The zippering of the neck region onto the head induces a conformation that allows hydrolysis of ATP (the head is an ATPase enzyme). This conformation (ADP bound, neck region zippered onto the head region) has lower affinity for the MT.
- The head releases the MT leading to a conformational change that causes the neck to unzipper from the head group. This restores the original ADP bound head domain with a high affinity for the MT. This head domain is ready for the next step.
- The hydrolysis of ATP has a very negative delta G so the reaction does not reverse.
Here is an animation of kinesin movement, Youtube version of Kinesin movement movie, Disco Kinesin totes a vesicle.
Here is an animation of myosin movement on actin, Youtube version of myosin movement movie. It uses a somewhat different mechanism even though structural studies now show that the kinesin and myosin are related and probably derive from the same ancestral protein.
Here is an animation of many different cellular processes that you now might be able to see are produced by application of simple biochemical principles. It can take a minute or so to load so be patient.
Charles S. Gasser (Department of Molecular & Cellular Biology; UC Davis)