Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.6.4.4 (kinesin)
5,033 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The actin cores of hair-cell stereocilia were tested as a substrate for the movement of myosin-coated beads in an in vitro assay. Large numbers of stereocilia from bullfrog sacculi and semicircular canals were isolated by blotting onto coverglasses and were demembranated to expose the polar actin tracks of their cytoskeletal cores. Silica or polystyrene beads, coated with thick filaments of chicken skeletal muscle myosin, were added to this core preparation in the presence of ATP. Myosin-coated beads could reach some of the cores by diffusion alone, but the efficiency and precision of the assay were improved considerably by the use of "optical tweezers" (a gradient-force optical trap) to deposit the beads directly on the cores. Beads applied in this fashion bound and moved unidirectionally at 1-2 microns/s, escaping the retarding force of the trap. Actin filaments within the stereocilia are cross-linked by fimbrin, but this did not appear to interfere with the motility of myosin. Beads coated with optic-lobe kinesin were also tested for movement; these bound and moved unidirectionally at 0.1-0.2 microns/s when applied to microtubule-based kinociliary cores, but not when applied to actin-based stereociliary cores. Our results are consistent with, and lend support to, a model for hair cell adaptation in which a molecular motor such as myosin maintains tension on the mechanically gated transduction channels. Optical tweezers and video-enhanced differential interference contrast optics provide high efficiency and improved optical resolution for the in vitro analysis of myosin motility.
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PMID:Actin cores of hair-cell stereocilia support myosin motility. 223 74

We show that microtubule polymers can be immobilized selectively on lithographically patterned silane surfaces while retaining their native properties. Silane films were chemisorbed on polished silicon wafers or glass coverslips and patterned using a deep UV lithographic process developed at the Naval Research Laboratory. Hydrocarbon and fluorocarbon alkyl silanes, as well as amino and thiol terminal alkyl silanes, were investigated as substrates for microtubule adhesion with retention of biological activity. Microtubules were found to adhere strongly to amine terminal silanes while retaining the ability to act as substrates for the molecular motor protein kinesin. Aminosilane patterns with linewidths varying from 1 to 50 microns were produced lithographically and used to produce patterns of selectively adhered microtubules. Microtubules were partially aligned on the patterned lines by performing the immobilization in a fluid flow field. Patterns were imaged with atomic force microscopy and differential interference contrast microscopy. Motility assays were carried out using kinesin-coated beads and observed with differential interference contrast microscopy. Kinesin bead movement on the patterned microtubules was comparable to movement on microtubule control surfaces.
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PMID:Selective adhesion of functional microtubules to patterned silane surfaces. 859 84

We demonstrate highly efficient rectification of microtubule motility on gold nanofabricated structures. First, we present a novel nanofabrication process for the creation of gold tracks for microtubule motility recessed in silicon oxide. This approach is particularly useful because it enables the use of the well-understood PEG-silane chemistry on SiO2 for the blocking of kinesin, whereas the gold tracks allow possible electrical control. We demonstrate excellent confinement of microtubule motility to the gold nanostructures and that microtubules move on the gold with speeds comparable to that on glass. Second, we present designs of three advanced rectifier geometries. We analyze the microtubule pathways through the geometries, and we demonstrate highly efficient rectification with up to 92% efficiency. As a result, we find that up to 97% of the microtubules move unidirectionally.
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PMID:High rectifying efficiencies of microtubule motility on kinesin-coated gold nanostructures. 1594 53

We report on a novel approach for the size-dependent fractionation of protein assemblies on polymeric surfaces. Using a simple temperature gradient method to generate one-dimensional gradients of grafted poly(ethylene glycol), we fabricated silicon-oxide chips with a gradually changing surface density of kinesin motor molecules. We demonstrate that such a bioactive surface can be used to sort gliding microtubules according to their length. To our knowledge, this is the first example of the self-organized sorting of protein assemblies on surfaces.
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PMID:Size sorting of protein assemblies using polymeric gradient surfaces. 1621 8

We report a novel approach for the dynamic control of gliding microtubule motility by external stimuli. Our approach is based on the fabrication of a composite surface where functional kinesin motor-molecules are adsorbed onto a silicon substrate between surface-grafted polymer chains of thermoresponsive poly(N-isopropylacrylamide). By external temperature control between 27 and 35 degrees C, we demonstrate the reversible landing, gliding, and releasing of motor-driven microtubules in response to conformational changes of the polymer chains. Our method represents a versatile means to control the activity of biomolecular motors, and other surface-coupled enzyme systems, in bionanotechnological applications.
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PMID:Reversible switching of microtubule motility using thermoresponsive polymer surfaces. 1696 12

Biosensors can be miniaturized by either injecting smaller volumes into micro- and nanofluidic devices or immersing increasingly sophisticated particles known as 'smart dust' into the sample. The term 'smart dust' originally referred to cubic-millimetre wireless semiconducting sensor devices that could invisibly monitor the environment in buildings and public spaces, but later it also came to include functional micrometre-sized porous silicon particles used to monitor yet smaller environments. The principal challenge in designing smart dust biosensors is integrating transport functions with energy supply into the device. Here, we report a hybrid microdevice that is powered by ATP and relies on antibody-functionalized microtubules and kinesin motors to transport the target analyte into a detection region. The transport step replaces the wash step in traditional double-antibody sandwich assays. Owing to their small size and autonomous function, we envision that large numbers of such smart dust biosensors could be inserted into organisms or distributed into the environment for remote sensing.
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PMID:A smart dust biosensor powered by kinesin motors. 1926 45

An alternative method of micro/nano-transport has been achieved by using motor proteins. Microtubules on a kinesin-coated surface have potential to act as a nano-transport system. When microtubules are used as carriers, either cargo or cargo linkers are attached on the microtubule surface. Such cargo attachments can significantly affect kinesin motion. To deal with the difficulty caused by molecular attachment to the microtubule surface, the cargo loading and transport mechanism should be separated. In this work, we propose to use micromachined needles as cargo carriers which then can be transported on microtubules. Because of the separation of needle functionalization and transport mechanism, functionalization of the needles can proceed without any effect on the microtubule structure, significantly increasing the possible types of cargo. We have fabricated silicon needles in mass numbers using a simple and effective method and have shown that the microtubule-needle composites are transported without affecting the kinesin activity.
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PMID:A nano-needle/microtubule composite gliding on a kinesin-coated surface for target molecule transport. 2002 55

Motor proteins function in in vivo ensembles to achieve cargo transport, flagellum motion, and mitotic cell division. Although the cooperativity of multiple motors is indispensable for physiological function, reconstituting the arrangement of motors in vitro is challenging, so detailed analysis of the functions of motor ensembles has not yet been achieved. Here, we developed an assay platform to study the motility of microtubules driven by a defined number of kinesin motors spaced in a definite manner. Gold (Au) nano-pillar arrays were fabricated on a silicon/silicon dioxide (Si/SiO₂) substrate with spacings of 100 nm to 500 nm. The thiol-polyethylene glycol (PEG)-biotin self-assembled monolayer (SAM) and silane-PEG-CH₃ SAM were then selectively formed on the pillars and SiO₂ surface, respectively. This allowed for both immobilization of kinesin molecules on Au nano-pillars in a precise manner and repulsion of kinesins from the SiO₂ surface. Using arrayed kinesin motors, we report that motor number and spacing do not influence the motility of microtubules driven by kinesin-1 motors. This assay platform is applicable to all kinds of biotinylated motors, allows the study of the effects of motor number and spacing, and is expected to reveal novel behaviors of motor proteins.
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PMID:Transport of microtubules according to the number and spacing of kinesin motors on gold nano-pillars. 3088 73

The guided gliding of cytoskeletal filaments, driven by biomolecular motors on nano/microstructured chips, enables novel applications in biosensing and biocomputation. However, expensive and time-consuming chip production hampers the developments. It is therefore important to establish protocols to regenerate the chips, preferably without the need to dismantle the assembled microfluidic devices which contain the structured chips. We here describe a novel method toward this end. Specifically, we use the small, nonselective proteolytic enzyme, proteinase K to cleave all surface-adsorbed proteins, including myosin and kinesin motors. Subsequently, we apply a detergent (5% SDS or 0.05% Triton X100) to remove the protein remnants. After this procedure, fresh motor proteins and filaments can be added for new experiments. Both, silanized glass surfaces for actin-myosin motility and pure glass surfaces for microtubule-kinesin motility were repeatedly regenerated using this approach. Moreover, we demonstrate the applicability of the method for the regeneration of nano/microstructured silicon-based chips with selectively functionalized areas for supporting or suppressing gliding motility for both motor systems. The results substantiate the versatility and a promising broad use of the method for regenerating a wide range of protein-based nano/microdevices.
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PMID:Regeneration of Assembled, Molecular-Motor-Based Bionanodevices. 3151 80

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