Biophysics of Molecular Motors
Our present work aims at understandig the structural basis of auto-inhibition in heteromeric kinesin-2. Is the previously identified asymmetric regulation of the heteromeric kinesin-2 KLP11/KLP20 a universally conserved mechanism in the kinesin-2 class? To address this question, we use enzymatic bulk assays and single molecule assays to investigate auto-inhibition of kinesin-2 motors from various organisms.
Torque generation of kinesin.
In long-range transport of cargo, prototypical kinesin-1 steps along a single protofilament on the microtubule, an astonishing behavior given the number of theoretically available binding sites on adjacent protofilaments. Using a laser trap assay, we analyzed the trajectories of several representatives from the kinesin-2 class on freely suspended microtubules. In stark contrast to kinesin-1, these motors display a wide range of left-handed spiraling around microtubules and thus generate torque during cargo transport. We provide direct evidence that kinesin's neck region determines the torque-generating properties and present a model that explains the physical basis of kinesin's spiraling around the microtubule. download
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How kinesin-2 forms a stalk.
The heterotrimeric structure of kinesin-2 makes it a unique member of the kinesin superfamily; however, molecular details of the oligomer formation are largely unknown. We demonstrate that heterodimerization of the two distinct motor domains KLP11 and KLP20 of Caenorhabditis elegans kinesin-2 requires a dimerization seed of merely two heptads at the C terminus of the stalk. This heterodimeric seed is sufficient to promote dimerization along the entire length of the stalk, as shown by circular dichroism spectroscopy, Förster resonance energy transfer analysis, and electron microscopy. In addition to explaining the formation of the kinesin-2 stalk, the seed sequence identified bears great potential for generating specific heterodimerization in other protein biochemical applications. pdf
Myosin VI walks hand-over-hand
Myosin VI is a molecular motor that can walk processively on actin filaments with a 36-nm step size. The walking mechanism of myosin VI is controversial because it takes very large steps without an apparent lever arm of required length. Therefore, myosin VI is argued to be the first exception to the widely established lever arm theory. It is therefore critical to directly demonstrate whether this motor walks hand-over-hand along actin despite its short lever arm. Here, we follow the displacement of a single myosin VI head during the stepping process. A single head is displaced 72 nm during stepping, whereas the center of mass previously has been shown to move 36 nm. The most likely explanation for this result is a hand-over-hand walking mechanism. We hypothesize the existence of a flexible element that would allow the motor to bridge the observed 72-nm distance. download