ORCID Profile
0000-0002-0750-8515
Current Organisation
Technische Universität Dresden
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In Research Link Australia (RLA), "Research Topics" refer to ANZSRC FOR and SEO codes. These topics are either sourced from ANZSRC FOR and SEO codes listed in researchers' related grants or generated by a large language model (LLM) based on their publications.
Plant Cell and Molecular Biology | Plant Biology | Plant Physiology | Nanotechnology | Structural Biology (incl. Macromolecular Modelling) | Biological Physics | Nanoelectronics
Climate Change Adaptation Measures | Rice | Production of Biofuels (Biomass) | Expanding Knowledge in Technology |
Publisher: Wiley
Date: 28-07-2006
Abstract: We have developed a technique to manipulate bifunctional DNA molecules: One end is thiolated to bind to a patterned gold surface and the other end is biotinylated to bind to a microtubule gliding over a kinesin‐coated surface. We found that DNA molecules can be stretched and overstretched between the gold pads and the motile microtubules, and that they can form dynamic networks. This serves as a proof‐of‐principle that biological machineries can be used in vitro to accomplish the parallel formation of structured DNA templates that will have applications in biophysics and nanoelectronics.
Publisher: Wiley
Date: 2007
DOI: 10.1002/AR.20599
Abstract: Cells are composed of macromolecular structures of various sizes that act in idually or collectively to maintain their viability and perform their function within the organism. This review focuses on one structure, the microtubule, and one of the motor proteins that move along it, conventional kinesin (kinesin 1). Recent work on the cellular functions of kinesins, such as the organization of microtubules during cellular ision and the movement of the organelles and vesicles, offers insights into how biological motors might prove useful for organizing structures in engineered environments.
Publisher: Elsevier BV
Date: 07-2014
Publisher: EDP Sciences
Date: 12-2012
Publisher: The Optical Society
Date: 05-02-2013
DOI: 10.1364/OE.21.003523
Publisher: Elsevier BV
Date: 02-2003
DOI: 10.1016/S1097-2765(03)00049-2
Abstract: MCAK belongs to the Kin I subfamily of kinesin-related proteins, a unique group of motor proteins that are not motile but instead destabilize microtubules. We show that MCAK is an ATPase that catalytically depolymerizes microtubules by accelerating, 100-fold, the rate of dissociation of tubulin from microtubule ends. MCAK has one high-affinity binding site per protofilament end, which, when occupied, has both the depolymerase and ATPase activities. MCAK targets protofilament ends very rapidly (on-rate 54 micro M(-1).s(-1)), perhaps by diffusion along the microtubule lattice, and, once there, removes approximately 20 tubulin dimers at a rate of 1 s(-1). We propose that up to 14 MCAK dimers assemble at the end of a microtubule to form an ATP-hydrolyzing complex that processively depolymerizes the microtubule.
Publisher: Elsevier BV
Date: 09-2009
DOI: 10.1016/J.CELL.2009.07.032
Abstract: Motor proteins in the kinesin-8 family depolymerize microtubules in a length-dependent manner that may be crucial for controlling the length of organelles such as the mitotic spindle. We used single-molecule microscopy to understand the mechanism of length-dependent depolymerization by the budding yeast kinesin-8, Kip3p. We found that after binding at a random position on a microtubule and walking to the plus end, an in idual Kip3p molecule pauses there until an incoming Kip3p molecule bumps it off. Kip3p dissociation is accompanied by removal of just one or two tubulin dimers (on average). Such a cooperative mechanism leads to a depolymerization rate that is proportional to the flux of motors to the microtubule end and accounts for the length dependence of depolymerization. This type of feedback between length and disassembly may serve as a model for understanding how an ensemble of molecules can measure and control polymer length.
Publisher: Proceedings of the National Academy of Sciences
Date: 24-10-2006
Abstract: Kinesin-1 is a motor protein that carries cellular cargo such as membrane-bounded organelles along microtubules (MTs). The homodimeric motor molecule contains two N-terminal motor domains (the motor “heads”), a long coiled-coil domain (the “rod” or “stalk”), and two small globular “tail” domains. Much has been learned about how kinesin's heads step along a MT and how the tail is involved in cargo binding and autoinhibition. However, little is known about the role of the rod. Here, we investigate the extension of the rod during active transport by measuring the height at which MTs glide over a kinesin-coated surface in the presence of ATP. To perform height measurements with nanometer precision, we used fluorescence interference contrast microscopy, which is based on the self-interference of fluorescent light from objects near a reflecting surface. Using an in situ calibrating method, we determined that kinesin-1 molecules elevate gliding MTs 17 ± 2 nm (mean ± SEM) above the surface. When varying the composition of the surrounding nucleotides or removing the negatively charged -COOH termini of the MTs by subtilisin digestion, we found no significant changes in the measured distance. Even though this distance is significantly shorter than the contour length of the motor molecule (≈60 nm), it may be sufficient to prevent proteins bound to the MTs or prevent the organelles from interfering with transport.
Publisher: IOP Publishing
Date: 06-2021
Abstract: Molecular motor gliding motility assays based on myosin/actin or kinesin/microtubules are of interest for nanotechnology applications ranging from cargo-trafficking in lab-on-a-chip devices to novel biocomputation strategies. Prototype systems are typically monitored by expensive and bulky fluorescence microscopy systems. The development of integrated, direct electric detection of single filaments would strongly benefit applications and scale-up. We present estimates for the viability of such a detector by calculating the electrostatic potential change generated at a carbon nanotube transistor by a motile actin filament or microtubule under realistic gliding assay conditions. We combine this with detection limits based on previous state-of-the-art experiments using carbon nanotube transistors to detect catalysis by a bound lysozyme molecule and melting of a bound short-strand DNA molecule. Our results show that detection should be possible for both actin and microtubules using existing low ionic strength buffers given good device design, e.g., by raising the transistor slightly above the guiding channel floor. We perform studies as a function of buffer ionic strength, height of the transistor above the guiding channel floor, presence/absence of the casein surface passivation layer for microtubule assays and the linear charge density of the actin filaments/microtubules. We show that detection of microtubules is a more likely prospect given their smaller height of travel above the surface, higher negative charge density and the casein passivation, and may possibly be achieved with the nanoscale transistor sitting directly on the guiding channel floor.
Publisher: Proceedings of the National Academy of Sciences
Date: 26-06-2007
Abstract: The stepping behavior of single kinesin-1 motor proteins has been studied in great detail. However, in cells, these motors often do not work alone but rather function in small groups when they transport cellular cargo. Until now, the cooperative interactions between motors in such groups were poorly understood. A fundamental question is whether two or more motors that move the same cargo step in synchrony, producing the same step size as a single motor, or whether the step size of the cargo movement varies. To answer this question, we performed in vitro gliding motility assays, where microtubules coated with quantum dots were driven over a glass surface by a known number of kinesin-1 motors. The motion of in idual microtubules was then tracked with nanometer precision. In the case of transport by two kinesin-1 motors, we found successive 4-nm steps, corresponding to half the step size of a single motor. Dwell-time analysis did not reveal any coordination, in the sense of alternate stepping, between the motors. When three motors interacted in collective transport, we identified distinct forward and backward jumps on the order of 10 nm. The existence of the fractional steps as well as the distinct jumps illustrate a lack of synchronization and has implications for the analysis of motor-driven organelle movement investigated in vivo .
Publisher: Elsevier
Date: 2010
Publisher: Proceedings of the National Academy of Sciences
Date: 19-03-2012
Abstract: Despite the crowdedness of the interior of cells, microtubule-based motor proteins are able to deliver cargoes rapidly and reliably throughout the cytoplasm. We hypothesize that motor proteins may be adapted to operate in crowded environments by having molecular properties that prevent them from forming traffic jams. To test this hypothesis, we reconstituted high-density traffic of purified kinesin-8 motor protein, a highly processive motor with long end-residency time, along microtubules in a total internal-reflection fluorescence microscopy assay. We found that traffic jams, characterized by an abrupt increase in the density of motors with an associated abrupt decrease in motor speed, form even in the absence of other obstructing proteins. To determine the molecular properties that lead to jamming, we altered the concentration of motors, their processivity, and their rate of dissociation from microtubule ends. Traffic jams occurred when the motor density exceeded a critical value (density-induced jams) or when motor dissociation from the microtubule ends was so slow that it resulted in a pileup (bottleneck-induced jams). Through comparison of our experimental results with theoretical models and stochastic simulations, we characterized in detail under which conditions density- and bottleneck-induced traffic jams form or do not form. Our results indicate that transport kinesins, such as kinesin-1, may be evolutionarily adapted to avoid the formation of traffic jams by moving only with moderate processivity and dissociating rapidly from microtubule ends.
Publisher: Elsevier BV
Date: 07-2012
Publisher: Cold Spring Harbor Laboratory
Date: 12-08-2020
DOI: 10.1101/2020.08.11.245910
Abstract: Cytoskeletal motors transform chemical energy into mechanical work to drive essential cellular functions. Optical trapping experiments have provided crucial insights into the operation of these molecular machines under load. However, the throughput of such force spectroscopy experiments is typically limited to one measurement at a time. Here, we introduce a highly-parallel, microfluidics-based method that allows for rapid collection of force-dependent motility parameters of cytoskeletal motors with two orders of magnitude improvement in throughput compared to currently available methods. We apply tunable hydrodynamic forces to stepping kinesin-1 motors via DNA-tethered beads and utilize a large field of view to simultaneously track the velocities, run lengths and interaction times of hundreds of in idual kinesin-1 molecules under varying resisting and assisting loads. Importantly, the 16-μm long DNA tethers between the motors and the beads significantly reduces the vertical component of the applied force pulling the motors away from the microtubule. Our approach is readily applicable to other molecular systems and constitutes a new methodology for parallelized single-molecule force studies on cytoskeletal motors.
Publisher: Wiley
Date: 23-03-2021
Publisher: Wiley
Date: 28-10-2022
DOI: 10.1111/JPY.13287
Abstract: Diatoms are single-celled microalgae with silica-based cell walls (frustules) that are abundantly present in aquatic habitats, and form the basis of the food chain in many ecosystems. Many benthic diatoms have the remarkable ability to glide on all natural or man-made underwater surfaces using a carbohydrate- and protein-based adhesive to generate traction. Previously, three glycoproteins, termed FACs (Frustule Associated Components), have been identified from the common fouling diatom Craspedostauros australis and were implicated in surface adhesion through inhibition studies with a glycan-specific antibody. The polypeptide sequences of FACs remained unknown, and it was unresolved whether the FAC glycoproteins are indeed involved in adhesion, or whether this is achieved by different components sharing the same glycan epitope with FACs. Here we have determined the polypeptide sequences of FACs using peptide mapping by LC-MS/MS. Unexpectedly, FACs share the same polypeptide backbone (termed CaFAP1), which has a domain structure of alternating Cys-rich and Pro-Thr/Ser-rich regions reminiscent of the gel-forming mucins. By developing a genetic transformation system for C. australis, we were able to directly investigate the function of CaFAP1-based glycoproteins in vivo. GFP-tagging of CaFAP1 revealed that it constitutes a coat around all parts of the frustule and is not an integral component of the adhesive. CaFAP1-GFP producing transformants exhibited the same properties as wild type cells regarding surface adhesion and motility speed. Our results demonstrate that FAC glycoproteins are not involved in adhesion and motility, but might rather act as a lubricant to prevent fouling of the diatom surface.
Publisher: Springer Science and Business Media LLC
Date: 05-2006
DOI: 10.1038/NATURE04736
Abstract: The microtubule cytoskeleton is a dynamic structure in which the lengths of the microtubules are tightly regulated. One regulatory mechanism is the depolymerization of microtubules by motor proteins in the kinesin-13 family. These proteins are crucial for the control of microtubule length in cell ision, neuronal development and interphase microtubule dynamics. The mechanism by which kinesin-13 proteins depolymerize microtubules is poorly understood. A central question is how these proteins target to microtubule ends at rates exceeding those of standard enzyme-substrate kinetics. To address this question we developed a single-molecule microscopy assay for MCAK, the founding member of the kinesin-13 family. Here we show that MCAK moves along the microtubule lattice in a one-dimensional (1D) random walk. MCAK-microtubule interactions were transient: the average MCAK molecule diffused for 0.83 s with a diffusion coefficient of 0.38 microm2 s(-1). Although the catalytic depolymerization by MCAK requires the hydrolysis of ATP, we found that the diffusion did not. The transient transition from three-dimensional diffusion to 1D diffusion corresponds to a "reduction in dimensionality" that has been proposed as the search strategy by which DNA enzymes find specific binding sites. We show that MCAK uses this strategy to target to both microtubule ends more rapidly than direct binding from solution.
Publisher: American Chemical Society (ACS)
Date: 22-08-2003
DOI: 10.1021/NL034504H
Publisher: Elsevier BV
Date: 05-2015
Publisher: Elsevier BV
Date: 03-2002
DOI: 10.1016/S0960-9822(02)00750-9
Abstract: A fusion protein of kinesin and gelsolin binds a short actin filament which can be visualized using a standard fluorescence microscope. This technique has provided new insight into the mechanism of kinesin action, and in principle it can be extended to allow single-molecule assays of any protein.
Start Date: 05-2021
End Date: 05-2025
Amount: $450,000.00
Funder: Australian Research Council
View Funded ActivityStart Date: 06-2019
End Date: 12-2021
Amount: $531,000.00
Funder: Australian Research Council
View Funded Activity