title: Watching cellular machinery in action, one molecule at a time.
abstract: Single-molecule manipulation and imaging techniques have become important elements of the biologist's toolkit to gain mechanistic insights into cellular processes. By removing ensemble averaging, single-molecule methods provide unique access to the dynamic behavior of biomolecules. Recently, the use of these approaches has expanded to the study of complex multiprotein systems and has enabled detailed characterization of the behavior of individual molecules inside living cells. In this review, we provide an overview of the various force- and fluorescence-based single-molecule methods with applications both in vitro and in vivo, highlighting these advances by describing their applications in studies on cytoskeletal motors and DNA replication. We also discuss how single-molecule approaches have increased our understanding of the dynamic behavior of complex multiprotein systems. These methods have shown that the behavior of multicomponent protein complexes is highly stochastic and less linear and deterministic than previously thought. Further development of single-molecule tools will help to elucidate the molecular dynamics of these complex systems both inside the cell and in solutions with purified components.
authors.affiliation: Centre for Medical and Molecular Bioscience, Illawarra Health and Medical Research Institute and University of Wollongong, New South Wales 2522, Australia.|Zernike Institute for Advanced Materials, University of Groningen, Groningen 9747 AG, Netherlands.
authors.forename: Enrico
authors.identifier: http://orcid.org/0000-0002-2032-2950
authors.initials: E
authors.lastname: Monachino
authors.affiliation: Centre for Medical and Molecular Bioscience, Illawarra Health and Medical Research Institute and University of Wollongong, New South Wales 2522, Australia.|Zernike Institute for Advanced Materials, University of Groningen, Groningen 9747 AG, Netherlands.
authors.forename: Lisanne M
authors.initials: LM
authors.lastname: Spenkelink
authors.affiliation: Centre for Medical and Molecular Bioscience, Illawarra Health and Medical Research Institute and University of Wollongong, New South Wales 2522, Australia vanoijen@uow.edu.au.
authors.forename: Antoine M
authors.initials: AM
authors.lastname: van Oijen
mesh_terms: D000818:Animals; D015374:Biosensing Techniques; D003585:Cell Biology; D003584:Cytological Techniques; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D004261:DNA Replication; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D008280:Magnetics; D018625:Microscopy, Atomic Force; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D057054:Molecular Imaging / Q000379:methods*; D020409:Molecular Motor Proteins / Q000378:metabolism*; D052898:Optical Tweezers
chemical_list: D003598:Cytoskeletal Proteins; D020409:Molecular Motor Proteins