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  • 1.
    Albet-Torres, Nuria
    et al.
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Gunnarsson, Anders
    Persson, Malin
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Balaz, Martina
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Höök, Fredrik
    Månsson, Alf
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Molecular motors on lipid bilayers and silicon dioxide: different driving forces for adsorption2010In: Soft Matter, ISSN 1744-683X, E-ISSN 1744-6848, Vol. 6, no 14, p. 3211-3219Article in journal (Refereed)
    Abstract [en]

    Understanding how different types of interactions govern adsorption of the myosin motor fragment heavy meromyosin (HMM) onto different substrates is important in functional studies of actomyosin and for the development of motor powered lab-on-a-chip applications. In this study, we have combined in vitro motility assays and quartz crystal microbalance with dissipation (QCM-D) monitoring to investigate the underlying adsorption mechanisms of HMM onto supported lipid bilayers in comparison with pure and silanized SiO2. The QCM-D results, combined with data showing actin transportation by HMM adsorbed onto positively charged supported lipid bilayers, suggest reversible HMM surface adsorption via the negatively charged coiled-coil tail region. In contrast, the QCM-D data for HMM adsorption onto negatively charged lipids support a model according to which HMM adsorbs onto negatively charged surfaces largely via the positively charged actin binding regions. Adsorption studies at low (30-65 mM) and high (185-245 mM) ionic strengths onto piranha cleaned SiO2 surfaces (contact angle < 20 degrees) support this general model. However, unlike the situation for charged lipids, rinsing in high ionic strength solution caused only partial HMM desorption from SiO2, without restoration of actin propulsion by the remaining HMM molecules. This suggests that mechanisms other than electrostatic interactions are involved in the tethering of HMM heads to SiO2 surfaces. An expanded model for HMM adsorption is formulated on the basis of the data and the potential of the results for nanotechnological applications of actomyosin is discussed.

  • 2. Aveyard,, J
    et al.
    Hajne, J.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    van Delft,, F.C.M.J.M.
    van Zijl, J.
    Snijder, J.
    van den Heuvel,, F.C.
    Nicolau,, D.V.
    Actin motility confinement on micro/nanostructured surfaces.2013In: Proc. SPI 8587, Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues XI, 858722 (February 22, 2013) / [ed] Daniel L. Farkas; Dan V. Nicolau; Robert C. Leif, SPIE - International Society for Optical Engineering, 2013, p. 858722-858727Conference paper (Refereed)
  • 3.
    Balaz, Martina
    et al.
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Sundberg, Mark
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Persson, Malin
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Kvassman, Jan-Olov
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Månsson, Alf
    University of Kalmar, School of Pure and Applied Natural Sciences.
    "Effects of surface adsorption on catalytic activity of heavy meromyosin studied using fluorescent ATP analogue"2007In: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 24, p. 4917-4934Article in journal (Refereed)
  • 4.
    Bengtsson, Elina
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Kumar, Saroj
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Actomyosin Interactions and Different Structural States of Actin Filaments2013In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 104, no 2, p. 480A-481AArticle in journal (Other academic)
  • 5.
    Bengtsson, Elina
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Kumar, Saroj
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Altered Structural State of Actin Filaments Upon MYOSIN II Binding2015In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 108, no 2 Supplement 1, p. 299A-300A, article id 1499-PosArticle in journal (Other academic)
  • 6.
    Bengtsson, Elina
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Analysis of Flexural Rigidity of Actin Filaments Propelled by Surface Adsorbed Myosin Motors2013In: Cytoskeleton, ISSN 1949-3584, Vol. 70, no 11, p. 718-728Article in journal (Refereed)
    Abstract [en]

    Actin filaments are central components of the cytoskeleton and the contractile machinery of muscle. The filaments are known to exist in a range of conformational states presumably with different flexural rigidity and thereby different persistence lengths. Our results analyze the approaches proposed previously to measure the persistence length from the statistics of the winding paths of actin filaments that are propelled by surface-adsorbed myosin motor fragments in the in vitro motility assay. Our results suggest that the persistence length of heavy meromyosin propelled actin filaments can be estimated with high accuracy and reproducibility using this approach provided that: (1) the in vitro motility assay experiments are designed to prevent bias in filament sliding directions, (2) at least 200 independent filament paths are studied, (3) the ratio between the sliding distance between measurements and the camera pixel-size is between 4 and 12, (4) the sliding distances between measurements is less than 50% of the expected persistence length, and (5) an appropriate cut-off value is chosen to exclude abrupt large angular changes in sliding direction that are complications, e.g., due to the presence of rigor heads. If the above precautions are taken the described method should be a useful routine part of in vitro motility assays thus expanding the amount of information to be gained from these. (c) 2013 Wiley Periodicals, Inc.

  • 7.
    Bengtsson, Elina
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences. Karolinska Institutet ; McGill Univ, Canada.
    Rahman, Mohammad A.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Kumar, Saroj
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences. Delhi Technol Univ, India.
    Takatsuki, Hideyo
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Myosin-Induced Gliding Patterns at Varied [MgATP] Unveil a Dynamic Actin Filament2016In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 111, no 7, p. 1465-1477Article in journal (Refereed)
    Abstract [en]

    Actin filaments have key roles in cell motility but are generally claimed to be passive interaction partners in actin-myosin -based motion generation. Here, we present evidence against this static view based on an altered myosin-induced actin filament gliding pattern in an in vitro motility assay at varied [MgATP]. The statistics that characterize the degree of meandering of the actin filament paths suggest that for [MgATP] >= 0.25 mM, the flexural rigidity of heavy meromyosin (HMM)-propelled actin filaments is similar (without phalloidin) or slightly lower (with phalloidin) than that of HMM-free filaments observed in solution without surface tethering. When [MgATP] was reduced to <= 0.1 mM, the actin filament paths in the in vitro motility assay became appreciably more winding in both the presence and absence of phalloidin. This effect of lowered [MgATP] was qualitatively different from that seen when HMM was mixed with ATP-insensitive, N-ethylmaleimide-treated HMM (NEM-HMM; 25-30%). In particular, the addition of NEM-HMM increased a non-Gaussian tail in the path curvature distribution as well as the number of events in which different parts of an actin filament followed different paths. These effects were the opposite of those observed with reduced [MgATP]. Theoretical modeling suggests a 30-40% lowered flexural rigidity of the actin filaments at [MgATP] <= 0.1 mM and local bending of the filament front upon each myosin head attachment. Overall, the results fit with appreciable structural changes in the actin filament during actomyosin-based motion generation, and modulation of the actin filament mechanical properties by the dominating chemomechanical actomyosin state.

  • 8.
    Kumar, Saroj
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Milani, Gloria
    University of Padova, Italy.
    Takatsuki, Hideyo
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Lana, Tobia
    University of Padova, Italy.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Frasson, Chiara
    University of Padova, Italy.
    te Kronnie, Geertruy
    University of Padova, Italy.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Sensing protein antigen and microvesicle analytes using high-capacity biopolymer nano-carriers2016In: The Analyst, ISSN 0003-2654, E-ISSN 1364-5528, Vol. 141, no 3, p. 836-846Article in journal (Refereed)
    Abstract [en]

    Lab-on-a-chip systems with molecular motor driven transport of analytes attached to cytoskeletal filament shuttles (actin filaments, microtubules) circumvent challenges with nanoscale liquid transport. However, the filaments have limited cargo-carrying capacity and limitations either in transportation speed (microtubules) or control over motility direction (actin). To overcome these constraints we here report incorporation of covalently attached antibodies into self-propelled actin bundles (nanocarriers) formed by cross-linking antibody conjugated actin filaments viafascin, a natural actin-bundling protein. We demonstrate high maximum antigen binding activity and propulsion by surface adsorbed myosin motors. Analyte transport capacity is tested using both protein antigens and microvesicles, a novel class of diagnostic markers. Increased incubation concentration with protein antigen in the 0.1–100 nM range (1 min) reduces the fraction of motile bundles and their velocity but maximum transportation capacity of >1 antigen per nm of bundle length is feasible. At sub-nanomolar protein analyte concentration, motility is very well preserved opening for orders of magnitude improved limit of detection using motor driven concentration on nanoscale sensors. Microvesicle-complexing to monoclonal antibodies on the nanocarriers compromises motility but nanocarrier aggregation via microvesicles shows unique potential in label-free detection with the aggregates themselves as non-toxic reporter elements.

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  • 9.
    Kumar, Saroj
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    ten Siethoff, Lasse
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Albet-Torres, Nuria
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Magnetic capture from blood rescues molecular motor function in diagnostic nanodevices2013In: Journal of Nanobiotechnology, E-ISSN 1477-3155, Vol. 11, article id 14Article in journal (Refereed)
    Abstract [en]

    Background: Introduction of effective point-of-care devices for use in medical diagnostics is part of strategies to combat accelerating health-care costs. Molecular motor driven nanodevices have unique potentials in this regard due to unprecedented level of miniaturization and independence of external pumps. However motor function has been found to be inhibited by body fluids. Results: We report here that a unique procedure, combining separation steps that rely on antibody-antigen interactions, magnetic forces applied to magnetic nanoparticles (MPs) and the specificity of the actomyosin bond, can circumvent the deleterious effects of body fluids (e.g. blood serum). The procedure encompasses the following steps: (i) capture of analyte molecules from serum by MP-antibody conjugates, (ii) pelleting of MP-antibody-analyte complexes, using a magnetic field, followed by exchange of serum for optimized biological buffer, (iii) mixing of MP-antibody-analyte complexes with actin filaments conjugated with same polyclonal antibodies as the magnetic nanoparticles. This causes complex formation: MP-antibody-analyte-antibody-actin, and magnetic separation is used to enrich the complexes. Finally (iv) the complexes are introduced into a nanodevice for specific binding via actin filaments to surface adsorbed molecular motors (heavy meromyosin). The number of actin filaments bound to the motors in the latter step was significantly increased above the control value if protein analyte (50-60 nM) was present in serum (in step i) suggesting appreciable formation and enrichment of the MP-antibody-analyte-antibody-actin complexes. Furthermore, addition of ATP demonstrated maintained heavy meromyosin driven propulsion of actin filaments showing that the serum induced inhibition was alleviated. Detailed analysis of the procedure i-iv, using fluorescence microscopy and spectroscopy identified main targets for future optimization. Conclusion: The results demonstrate a promising approach for capturing analytes from serum for subsequent motor driven separation/detection. Indeed, the observed increase in actin filament number, in itself, signals the presence of analyte at clinically relevant nM concentration without the need for further motor driven concentration. Our analysis suggests that exchange of polyclonal for monoclonal antibodies would be a critical improvement, opening for a first clinically useful molecular motor driven lab-on-a-chip device.

  • 10.
    Kumar, Saroj
    et al.
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    ten Siethoff, Lasse
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Persson, Malin
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Lard, Mercy
    Lund University.
    Kronnie, Geertruy Te
    University of Padua, Italy.
    Linke, Heiner
    Lund University.
    Månsson, Alf
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Antibodies Covalently Immobilized on Actin Filaments for Fast Myosin Driven Analyte Transport2012In: PLOS ONE, E-ISSN 1932-6203, Vol. 7, no 10, article id e46298Article in journal (Refereed)
    Abstract [en]

    Biosensors would benefit from further miniaturization, increased detection rate and independence from external pumps and other bulky equipment. Whereas transportation systems built around molecular motors and cytoskeletal filaments hold significant promise in the latter regard, recent proof-of-principle devices based on the microtubule-kinesin motor system have not matched the speed of existing methods. An attractive solution to overcome this limitation would be the use of myosin driven propulsion of actin filaments which offers motility one order of magnitude faster than the kinesin-microtubule system. Here, we realized a necessary requirement for the use of the actomyosin system in biosensing devices, namely covalent attachment of antibodies to actin filaments using heterobifunctional cross-linkers. We also demonstrated consistent and rapid myosin II driven transport where velocity and the fraction of motile actin filaments was negligibly affected by the presence of antibody-antigen complexes at rather high density (>20 mu m(-1)). The results, however, also demonstrated that it was challenging to consistently achieve high density of functional antibodies along the actin filament, and optimization of the covalent coupling procedure to increase labeling density should be a major focus for future work. Despite the remaining challenges, the reported advances are important steps towards considerably faster nanoseparation than shown for previous molecular motor based devices, and enhanced miniaturization because of high bending flexibility of actin filaments.

  • 11.
    Lard, Mercy
    et al.
    Lund University.
    ten Siethoff, Lasse
    Linnaeus University, Faculty of Social Sciences, Department of Sport Science.
    Generosi, Johanna
    Lund University.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Linke, Heiner
    Lund University.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Nanowire-Imposed Geometrical Control in Studies of Actomyosin Motor Function2015In: IEEE Transactions on Nanobioscience, ISSN 1536-1241, E-ISSN 1558-2639, Vol. 14, no 3, p. 289-297Article in journal (Refereed)
    Abstract [en]

    Recently, molecular motor gliding assays with actin and myosin from muscle have been realized on semiconductor nanowires coated with Al2O3. This opens for unique nanotechnological applications and novel fundamental studies of actomyosin motor function. Here, we provide a comparison of myosin-driven actin filament motility on Al2O3 to both nitrocellulose and trimethylchlorosilane derivatized surfaces. We also show that actomyosin motility on the less than 200 nm wide tips of arrays of Al2O3-coated nanowires can be used to control the number, and density, of myosin-actin attachment points. Results obtained using nanowire arrays with different inter-wire spacing are consistent with the idea that the actin filament sliding velocity is determined both by the total number and the average density of attached myosin heads along the actin filament. Further, the results are consistent with buckling of long myosin-free segments of the filaments as a factor underlying reduced velocity. On the other hand, the findings do not support a mechanistic role in decreasing velocity, of increased nearest neighbor distance between available myosin heads. Our results open up for more advanced studies that may use nanowire-based structures for fundamental investigations of molecular motors, including the possibility to create a nanowire-templated bottom-up assembly of 3D, muscle-like structures.

  • 12.
    Lard, Mercy
    et al.
    Lund University.
    ten Siethoff, Lasse
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Kumar, Saroj
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences. Delhi Technological University, India.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    te Kronnie, G.
    University of Padova, Italy.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Linke, H.
    Lund University.
    Nano-structuring for molecular motor control2015In: Nano-Structures for Optics and Photonics: Optical Strategies for Enhancing Sensing, Imaging, Communication and Energy Conversion / [ed] Baldassare Di Bartolo, John Collins, Luciano Silvestri, Springer, 2015, p. 459-459Conference paper (Refereed)
    Abstract [en]

    The interaction of self-propelled biological molecular-motors and cytoskeletal filaments holds relevance for a variety of applications such as biosensing, drug screening, diagnostics and biocomputation. The use of these systems for lab-on-a-chip biotechnology applications shows potential for replacement of microfluidic flow by active, molecular-motor driven transport of filaments. The ability to control, confine and detect motile objects in such a system is possible by development of nanostructured surfaces for on-chip applications and fundamental studies of molecular-motors. Here we describe the localized detection (Lard et al., Sci Rep 3:1092, 2013) and fast transport of actin filaments by myosin molecular-motors (Lard et al., Biosens Biolectron 48(0):145–152, 2013), inserted within nanostructures, as a method for biocomputation and molecular concentration. These results include extensive myosin driven concentration of actin filaments on a miniaturized detector, of relevance for use of molecular-motors in a diagnostics platform. Also, we discuss the local enhancement of the fluorescence signal of filaments, relevant for use in a biocomputation device where tracking of potentially thousands of motile objects is of primary significance.

  • 13.
    Lard, Mercy
    et al.
    The Nanometer Structure Consortium (nmC@LU), Division of Solid State Physics, Lund University.
    ten Siethoff, Lasse
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Kumar, Saroj
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    te Kronnie, Geertruy
    Department of Women's and Children's Health, University of Padova.
    Linke, Heiner
    The Nanometer Structure Consortium (nmC@LU), Division of Solid State Physics, Lund University.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Ultrafast molecular motor driven nanoseparation and biosensing2013In: Biosensors & bioelectronics, ISSN 0956-5663, E-ISSN 1873-4235, Vol. 48, p. 145-152Article in journal (Refereed)
    Abstract [en]

    Portable biosensor systems would benefit from reduced dependency on external power supplies as well as from further miniaturization and increased detection rate. Systems built around self-propelled biological molecular motors and cytoskeletal filaments hold significant promise in these regards as they are built from nanoscale components that enable nanoseparation independent of fluidic pumping. Previously reported microtubule-kinesin based devices are slow, however, compared to several existing biosensor systems. Here we demonstrate that this speed limitation can be overcome by using the faster actomyosin motor system. Moreover, due to lower flexural rigidity of the actin filaments, smaller features can be achieved compared to microtubule-based systems, enabling further miniaturization. Using a device designed through optimization by Monte Carlo simulations, we demonstrate extensive myosin driven enrichment of actin filaments on a detector area of less than 10 μm2, with a concentration half-time of approximately 40 s. We also show accumulation of model analyte (streptavidin at nanomolar concentration in nanoliter effective volume) detecting increased fluorescence intensity within seconds after initiation of motor-driven transportation from capture regions. We discuss further optimizations of the system and incorporation into a complete biosensing workflow.

  • 14.
    Matusovsky, Oleg S.
    et al.
    McGill Univ, Canada.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences. Linnaeus University, Linnaeus Knowledge Environments, Advanced Materials.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences. McGill Univ, Canada.
    Cheng, Yu-Shu
    McGill Univ, Canada.
    Rassier, Dilson E.
    McGill Univ, Canada.
    High-speed AFM reveals subsecond dynamics of cardiac thin filaments upon Ca2+ activation and heavy meromyosin binding2019In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 116, no 33, p. 16384-16393Article in journal (Refereed)
    Abstract [en]

    High-speed atomic force microscopy (HS-AFM) can be used to study dynamic processes with real-time imaging of molecules within 1- to 5-nm spatial resolution. In the current study, we evaluated the 3-state model of activation of cardiac thin filaments (cTFs) isolated as a complex and deposited on a mica-supported lipid bilayer. We studied this complex for dynamic conformational changes 1) at low and high [Ca2+] (pCa 9.0 and 4.5), and 2) upon myosin binding to the cTF in the nucleotide-free state or in the presence of ATP. HS-AFM was used to directly visualize the tropo-myosin-troponin complex and Ca2+-induced tropomyosin movements accompanied by structural transitions of actin monomers within cTFs. Our data show that cTFs at relaxing or activating conditions are not ultimately in a blocked or activated state, respectively, but rather the combination of states with a prevalence that is dependent on the [Ca2+] and the presence of weakly or strongly bound myosin. The weakly and strongly bound myosin induce similar changes in the structure of cTFs as confirmed by the local dynamical displacement of individual tropomyosin strands in the center of a regulatory unit of cTF at the relaxed and activation conditions. The displacement of tropomyosin at the relaxed conditions had never been visualized directly and explains the ability of myosin binding to TF at the relaxed conditions. Based on the ratios of nonactivated and activated segments within cTFs, we proposed a mechanism of tropomyosin switching from different states that includes both weakly and strongly bound myosin.

  • 15.
    Månsson, Alf
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences. Linnaeus University, Linnaeus Knowledge Environments, Advanced Materials.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences. McGill Univ, Canada;Karolinska Institutet, Sweden.
    Shalabi, Nabil
    McGill Univ, Canada.
    Rassier, Dilson E.
    McGill Univ, Canada.
    Nonlinear Actomyosin Elasticity in Muscle?2019In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 116, no 2, p. 330-346Article in journal (Refereed)
    Abstract [en]

    Cyclic interactions between myosin II motor domains and actin filaments that are powered by turnover of ATP underlie muscle contraction and have key roles in motility of nonmuscle cells. The elastic characteristics of actin-myosin cross-bridges are central in the force-generating process, and disturbances in these properties may lead to disease. Although the prevailing paradigm is that the cross-bridge elasticity is linear (Hookean), recent single-molecule studies suggest otherwise. Despite convincing evidence for substantial nonlinearity of the cross-bridge elasticity in the single-molecule work, this finding has had limited influence on muscle physiology and physiology of other ordered cellular actin-myosin ensembles. Here, we use a biophysical modeling approach to close the gap between single molecules and physiology. The model is used for analysis of available experimental results in the light of possible nonlinearity of the cross-bridge elasticity. We consider results obtained both under rigor conditions (in the absence of ATP) and during active muscle contraction. Our results suggest that a wide range of experimental findings from mechanical experiments on muscle cells are consistent with nonlinear actin-myosin elasticity similar to that previously found in single molecules. Indeed, the introduction of nonlinear cross-bridge elasticity into the model improves the reproduction of key experimental results and eliminates the need for force dependence of the ATP-induced detachment rate, consistent with observations in other single-molecule studies. The findings have significant implications for the understanding of key features of actin-myosin-based production of force and motion in living cells, particularly in muscle, and for the interpretation of experimental results that rely on stiffness measurements on cells or myofibrils.

  • 16.
    Nicolau, Dan V., Jr.
    et al.
    Univ Calif Berkeley,USA ; Mol Sense Ltd, UK.
    Lard, Mercy
    Lund University.
    Korten, Till
    Tech Univ Dresden, Germany ; Max Planck Inst Mol Cell Biol & Genet, Germany.
    van Delft, Falco C. M. J. M.
    Philips Res MiPlaza & Philips Innovat Serv, Netherlands.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Bengtsson, Elina
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Diez, Stefan
    Tech Univ Dresden, Germany ; Max Planck Inst Mol Cell Biol & Genet, Germany.
    Linke, Heiner
    Lund University.
    Nicolau, Dan V.
    Univ Liverpool, UK ; McGill Univ, Canada.
    REPLY TO EINARSSON: The computational power of parallel network exploration with many bioagents2016In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 113, no 23, p. E3188-E3188Article in journal (Refereed)
  • 17.
    Nicolau, Dan V., Jr.
    et al.
    University of California, USA ; Molecular Sense, Ltd., UK.
    Lard, Mercy
    Lund University.
    Korten, Till
    Technische Universität Dresden, Germany ; Max Planck Institute of Molecular Cell Biology and Genetics, Germany.
    van Delftf, Falco C. M. J. M.
    Philips Research MiPlaza, The Netherlands ; Philips Innovation Services, The Netherlands.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Bengtsson, Elina
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Diez, Stefan
    Technische Universität Dresden, Germany ; Max Planck Institute of Molecular Cell Biology and Genetics, Germany.
    Linke, Heiner
    Lund University.
    Nicolau, Dan V.
    University of Liverpool, UK ; McGill University, Canada.
    Parallel computation with molecular-motor-propelled agents in nanofabricated networks2016In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 113, no 10, p. 2591-2596Article in journal (Refereed)
    Abstract [en]

    The combinatorial nature of many important mathematical problems, including nondeterministic-polynomial-time (NP)-complete problems, places a severe limitation on the problem size that can be solved with conventional, sequentially operating electronic computers. There have been significant efforts in conceiving parallel-computation approaches in the past, for example: DNA computation, quantum computation, and microfluidics-based computation. However, these approaches have not proven, so far, to be scalable and practical from a fabrication and operational perspective. Here, we report the foundations of an alternative parallel-computation system in which a given combinatorial problem is encoded into a graphical, modular network that is embedded in a nanofabricated planar device. Exploring the network in a parallel fashion using a large number of independent, molecular-motor-propelled agents then solves the mathematical problem. This approach uses orders of magnitude less energy than conventional computers, thus addressing issues related to power consumption and heat dissipation. We provide a proof-of-concept demonstration of such a device by solving, in a parallel fashion, the small instance {2, 5, 9} of the subset sum problem, which is a benchmark NP-complete problem. Finally, we discuss the technical advances necessary to make our system scalable with presently available technology.

  • 18.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Characterization and optimization of the in vitro motility assay for fundamental studies of myosin II2013Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Myosin II is the molecular motor responsible for muscle contraction. It transforms the chemical energy in ATP into mechanical work while interacting with actin filaments in so called cross-bridge cycles. Myosin II or its proteolytic fragments e.g., heavy meromyosin (HMM) can be adsorbed to moderately hydrophobic surfaces in vitro, while maintaining their ability to translocate actin filaments. This enables observation of myosin-induced actin filament sliding in a microscope. This “in vitro motility assay” (IVMA) is readily used in fundamental studies of actomyosin, including studies of muscle contraction. The degree of correlation of the myosin II function in the IVMA with its function in muscle depends on how the myosin molecules are arranged on the surface. Therefore a multi-technique approach, including total internal reflection spectroscopy, fluorescence interference contrast microscopy and quartz crystal microbalance with dissipation, was applied to characterize the HMM surface configurations. Several configurations with varying distributions were identified depending on the surface property. The most favorable HMM configurations for actin binding were observed on moderately hydrophobic surfaces.

    The effects on actomyosin function of different cargo sizes and amount of cargo loaded on an actin filament were also investigated. No difference in sliding velocities could be observed, independent of cargo size indicating that diffusional processive runs of myosin II along an actin filament are not crucial for actomyosin function in muscle. Furthermore, a tool for accurate velocity measurements appropriate for IVMAs at low [MgATP] was developed by utilizing the actin filament capping protein CapZ. These improvements allowed an investigation of the [MgATP]-velocity relationship to study possible processivity in fast skeletal muscle myosin II.  It is shown that the [MgATP]–velocity relationship is well described by a Michaelis-Menten hyperbola.  In addition, statistical cross-bridge modeling showed that the experimental results are in good agreement with recent findings of actomyosin cross-bridge properties, e.g., non-linear cross-bridge elasticity. However, no effect of inter-head cooperativity could be observed.

     In conclusion, the described results have contributed to in-depth understanding of the actomyosin cross-bridge cycle in muscle contraction.

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  • 19.
    Persson, Malin
    et al.
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Albet-Torres, Nuria
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Ionov, Leonid
    Sundberg, Mark
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Höök, Fredrik
    Diez, Stefan
    Månsson, Alf
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Balaz, Martina
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Heavy meromyosin molecules extending more than 50 nm above adsorbing electronegative surfaces.2010In: Langmuir, ISSN 0743-7463, E-ISSN 1520-5827, Vol. 26, no 12, p. 9927-9936Article in journal (Refereed)
    Abstract [en]

    In the in vitro motility assay, actin filaments are propelled by surface-adsorbed myosin motors, or rather, myosin motor fragments such as heavy meromyosin (HMM). Recently, efforts have been made to develop actomyosin powered nanodevices on the basis of this assay but such developments are hampered by limited understanding of the HMM adsorption geometry. Therefore, we here investigate the HMM adsorption geometries on trimethylchlorosilane- [TMCS-] derivatized hydrophobic surfaces and on hydrophilic negatively charged surfaces (SiO(2)). The TMCS surface is of great relevance in fundamental studies of actomyosin and both surface substrates are important for the development of motor powered nanodevices. Whereas both the TMCS and SiO(2) surfaces were nearly saturated with HMM (incubation at 120 microg mL(-1)) there was little actin binding on SiO(2) in the absence of ATP and no filament sliding in the presence of ATP. This contrasts with excellent actin-binding and motility on TMCS. Quartz crystal microbalance with dissipation (QCM-D) studies demonstrate a HMM layer with substantial protein mass up to 40 nm above the TMCS surface, considerably more than observed for myosin subfragment 1 (S1; 6 nm). Together with the excellent actin transportation on TMCS, this strongly suggests that HMM adsorbs to TMCS mainly via its most C-terminal tail part. Consistent with this idea, fluorescence interference contrast (FLIC) microscopy showed that actin filaments are held by HMM 38 +/- 2 nm above the TMCS-surface with the catalytic site, on average, 20-30 nm above the surface. Viewed in a context with FLIC, QCM-D and TIRF results, the lack of actin motility and the limited actin binding on SiO(2) shows that HMM adsorbs largely via the actin-binding region on this surface with the C-terminal coiled-coil tails extending >50 nm into solution. The results and new insights from this study are of value, not only for the development of motor powered nanodevices but also for the interpretation of fundamental biophysical studies of actomyosin function and for the understanding of surface-protein interactions in general.

  • 20.
    Persson, Malin
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Bengtsson, Elina
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    ten Siethoff, Lasse
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Nonlinear Cross-Bridge Elasticity and Post-Power-Stroke Events in Fast Skeletal Muscle Actomyosin2013In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 105, no 8, p. 1871-1881Article in journal (Refereed)
    Abstract [en]

    Generation-of force and movement by actomyosin cross-bridges is the molecular basis of muscle contraction, but generally accepted ideas about cross-bridge properties have recently been questioned. Of the utmost significance, evidence for nonlinear cross-bridge elasticity has been presented. We here investigate how this and other newly discovered or postulated phenomena would modify cross-bridge operation, with focus on post-power-stroke events. First, as an experimental basis, we present evidence for a hyperbolic [MgATP]-velocity relationship of heavy-meromyosin-propelled actin filaments in the in vitro motility assay using fast rabbit skeletal muscle myosin (28-29 degrees C). As the hyperbolic [MgATP]-velocity relationship was not consistent with interhead cooperativity, we developed a cross-bridge model with independent myosin heads and strain-dependent interstate transition rates. The model, implemented with inclusion of MgATP-independent detachment from the rigor state, as suggested by previous single-molecule mechanics experiments, accounts well for the [MgATP]-velocity relationship if nonlinear cross-bridge elasticity is assumed, but not if linear cross-bridge elasticity is assumed. In addition, a better fit is obtained with load-independent than with load-dependent MgATP-induced detachment rate. We discuss our results in relation to previous data showing a nonhyperbolic [MgATP1-velocity relationship when actin filaments are propelled by myosin subfragment 1 or full-length myosin. We also consider the implications of our results for characterization of the cross-bridge elasticity in the filament lattice of muscle.

  • 21.
    Persson, Malin
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Bengtsson, Elina
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    ten Siethoff, Lasse
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Non-Linear Cross-Bridge Elasticity, ATP-Independent Detachment and ATP-Velocity Relationships for Skeletal Muscle Actomyosin2014In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 106, no 2, p. 158A-158AArticle in journal (Other academic)
  • 22.
    Persson, Malin
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Gullberg, Maria
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Tolf, Conny
    Linnaeus University, Faculty of Health and Life Sciences, Department of Biology and Environmental Science.
    Lindberg, A. Michael
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Kocer, Armagan
    University of Groningen, The Netherlands.
    Transportation of Nanoscale Cargoes by Myosin Propelled Actin Filaments2013In: PLOS ONE, E-ISSN 1932-6203, Vol. 8, no 2, article id e55931Article in journal (Refereed)
    Abstract [en]

    Myosin II propelled actin filaments move ten times faster than kinesin driven microtubules and are thus attractive candidates as cargo-transporting shuttles in motor driven lab-on-a-chip devices. In addition, actomyosin-based transportation of nanoparticles is useful in various fundamental studies. However, it is poorly understood how actomyosin function is affected by different number of nanoscale cargoes, by cargo size, and by the mode of cargo-attachment to the actin filament. This is studied here using biotin/fluorophores, streptavidin, streptavidin-coated quantum dots, and liposomes as model cargoes attached to monomers along the actin filaments ("side-attached") or to the trailing filament end via the plus end capping protein CapZ. Long-distance transportation (> 100 mu m) could be seen for all cargoes independently of attachment mode but the fraction of motile filaments decreased with increasing number of side-attached cargoes, a reduction that occurred within a range of 10-50 streptavidin molecules, 1-10 quantum dots or with just 1 liposome. However, as observed by monitoring these motile filaments with the attached cargo, the velocity was little affected. This also applied for end-attached cargoes where the attachment was mediated by CapZ. The results with side-attached cargoes argue against certain models for chemomechanical energy transduction in actomyosin and give important insights of relevance for effective exploitation of actomyosin-based cargo-transportation in molecular diagnostics and other nanotechnological applications. The attachment of quantum dots via CapZ, without appreciable modulation of actomyosin function, is useful in fundamental studies as exemplified here by tracking with nanometer accuracy.

  • 23. Ramsey, L.C.
    et al.
    Aveyard, J.
    van Zalinge, H.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Nicolau, DV.
    Electric field modulation of the motility of actin filaments on myosin-functionalised surfaces.2013In: Proc. SPIE 8594, Nanoscale Imaging, Sensing, and Actuation for Biomedical Applications X, 85940R (February 19, 2013) / [ed] Alexander N. Cartwright; Dan V. Nicolau, SPIE - International Society for Optical Engineering, 2013, p. 85940R-1-85940R-7Conference paper (Refereed)
  • 24.
    Shalabi, Nabil
    et al.
    McGill Univ, Canada;McGill Univ, Canada.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences. McGill Univ, Canada;Karolinska Institutet.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Vengallatore, Srikar
    McGill Univ, Canada.
    Rassier, Dilson E.
    McGill Univ, Canada.
    Sarcomere Stiffness during Stretching and Shortening of Rigor Skeletal Myofibrils2017In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 113, no 12, p. 2768-2776Article in journal (Refereed)
    Abstract [en]

    In this study, we measured the stiffness of skeletal muscle myofibrils in rigor. Using a custom-built atomic force microscope, myofibrils were first placed in a rigor state then stretched and shortened at different displacements (0.1-0.3 mu m per sarcomere) and nominal speeds (0.4 and 0.8 mu m/s). During stretching, the myofibril stiffness was independent of both displacement and speed (average of 987 nN/mu m). During shortening, the myofibril stiffness was independent of displacement, but dependent on speed (1234 nN/mu m at 0.4 mu m/s; 1106 nN/mu m at 0.8 mu m/s). Furthermore, the myofibril stiffness during shortening was greater than that during stretching and the difference depended on speed (31 % at 0.4 mu m/s; 8% at 0.8 mu m/s). The results suggest that the myofibrils exhibit nonlinear viscoelastic properties that may be derived from myofibril filaments, similar to what has been observed in muscle fibers.

  • 25. van Zalinge, Harm
    et al.
    Aveyard, Jenny
    Hajne, Joanna
    Persson, Malin
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Månsson, Alf
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Nicolau, Dan V.
    Actin Filament Motility Induced Variation of Resonance Frequency and Rigidity of Polymer Surfaces Studied by Quartz Crystal Microbalance2012In: Langmuir, ISSN 0743-7463, E-ISSN 1520-5827, Vol. 28, no 42, p. 15033-15037Article in journal (Refereed)
    Abstract [en]

    This contribution reports on the quantification of the parameters of the motility assays for actomyosin system using a quartz crystal microbalance (QCM). In particular, we report on the difference in the observed resonance frequency and dissipation of a quartz crystal when actin filaments are stationary as opposed to when they are motile. The changes in QCM measurements were studied for various polymer-coated surfaces functionalized with heavy meromyosin (HMM). The results of the QCM experiments show that the HMM-induced sliding velocity of actin filaments is modulated by a combination of the viscoelastic properties of the polymer layer including the HMM motors.

  • 26.
    van Zalinge, Harm
    et al.
    Univ Liverpool, UK.
    Ramsey, Laurence C.
    Univ Liverpool, UK.
    Aveyard, Jenny
    Univ Liverpool, UK.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Nicolau, Dan V.
    Univ Liverpool, UK ; McGill Univ, Canada.
    Surface-Controlled Properties of Myosin Studied by Electric Field Modulation2015In: Langmuir, ISSN 0743-7463, E-ISSN 1520-5827, Vol. 31, no 30, p. 8354-8361Article in journal (Refereed)
    Abstract [en]

    The efficiency of dynamic nanodevices using surface-immobilized protein molecular motors, which have been proposed for diagnostics, drug discovery, and biocomputation, critically depends on the ability to precisely control the motion of motor-propelled, individual cytoskeletal filaments transporting cargo to designated locations. The efficiency of these devices also critically depends on the proper function of the propelling motors, which is controlled by their interaction with the surfaces they are immobilized on. Here we use a microfluidic device to study how the motion of the motile elements, i.e., actin filaments propelled by heavy mero-myosin (HMM) motor fragments immobilized on various surfaces, is altered by the application of electrical loads generated by an external electric field with strengths ranging from 0 to 8 kVm(-1). Because the motility is intimately linked to the function of surface-immobilized motors, the study also showed how the adsorption properties of HMM on various surfaces, such as nitrocellulose (NC), trimethylclorosilane (TMCS), poly(methyl methacrylate) (PMMA), poly(tert-butyl methacrylate) (PtBMA), and poly(butyl methacrylate) (PBMA), can be characterized using an external field. It was found that at an electric field of 5 kVm(-1) the force exerted on the filaments is sufficient to overcome the frictionlike resistive force of the inactive motors. It was also found that the effect of assisting electric fields on the relative increase in the sliding velocity was markedly higher for the TMCS-derivatized surface than for all other polymer-based surfaces. An explanation of this behavior, based on the molecular rigidity of the TMCS-on-glass surfaces as opposed to the flexibility of the polymer-based ones, is considered. To this end, the proposed microfluidic device could be used to select appropriate surfaces for future lab-on-a-chip applications as illustrated here for the almost ideal TMCS surface. Furthermore, the proposed methodology can be used to gain fundamental insights into the functioning of protein molecular motors, such as the force exerted by the motors under different operational conditions.

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