Nov 18, 2014

Volume 107Issue 10p2221-2478, L25-L32
Open Archive
Cover picture: Vibrio cholerae biofilm at the air-liquid interface. Bacteria secrete proteins, polysaccharides, and other components to assemble a spider-web-like matrix that surrounds cells to promote the formation of protective communities termed biofilms. Biofilm formation increases both the survival and infectivity of Vibrio cholerae, the causative agent of cholera. This scanning electron micrograph captures a mature V. cholerae biofilm formed at the air-liquid interface. Improved models of biofilm assembly, structure, and function are needed to drive molecular strategies to interfere with V. cholerae biofilm formation. In our contribution to this issue, we integrated interfacial rheology measurements, electron microscopy, and molecular profiling to define distinct contributions of matrix proteins to biofilm morphology, architecture, and mechanical properties and discovered that the Bap1 protein is uniquely important for the integrity of biofilm structure and the maintenance of biofilm strength at the air-liquid interface. For more information, see the article by Hollenbeck et al. on page 2245....
Cover picture: Vibrio cholerae biofilm at the air-liquid interface. Bacteria secrete proteins, polysaccharides, and other components to assemble a spider-web-like matrix that surrounds cells to promote the formation of protective communities termed biofilms. Biofilm formation increases both the survival and infectivity of Vibrio cholerae, the causative agent of cholera. This scanning electron micrograph captures a mature V. cholerae biofilm formed at the air-liquid interface. Improved models of biofilm assembly, structure, and function are needed to drive molecular strategies to interfere with V. cholerae biofilm formation. In our contribution to this issue, we integrated interfacial rheology measurements, electron microscopy, and molecular profiling to define distinct contributions of matrix proteins to biofilm morphology, architecture, and mechanical properties and discovered that the Bap1 protein is uniquely important for the integrity of biofilm structure and the maintenance of biofilm strength at the air-liquid interface. For more information, see the article by Hollenbeck et al. on page 2245.

Biophysical Letters

  • Biophysical Letter

    Voltage Sensor Gating Charge Transfer in a hERG Potassium Channel Model

    • Charlotte K. Colenso,
    • Yang Cao,
    • Richard B. Sessions,
    • Jules C. Hancox,
    • Christopher E. Dempsey
    Relaxation of a hERG K+ channel model during molecular-dynamics simulation in a hydrated POPC bilayer was accompanied by transitions of an arginine gating charge across a charge transfer center in two voltage sensor domains. Inspection of the passage of arginine side chains across the charge transfer center suggests that the unique hydration properties of the arginine guanidine cation facilitates charge transfer during voltage sensor responses to changes in membrane potential, and underlies the preference of Arg over Lys as a mobile charge carrier in voltage-sensitive ion channels.
  • Biophysical Letter

    Deactivation of a Negative Regulator: A Distinct Signal Transduction Mechanism, Pronounced in Akt Signaling

    • Anisur Rahman,
    • Jason M. Haugh
    Kinase cascades, in which enzymes are sequentially activated by phosphorylation, are quintessential signaling pathways. Signal transduction is not always achieved by direct activation, however. Often, kinases activate pathways by deactivation of a negative regulator; this indirect mechanism, pervasive in Akt signaling, has yet to be systematically explored. Here, we show that the indirect mechanism has properties that are distinct from direct activation. With comparable parameters, the indirect mechanism yields a broader range of sensitivity to the input, beyond saturation of regulator phosphorylation, and kinetics that become progressively slower, not faster, with increasing input strength.

Cell Biophysics

  • Article

    Raman and Autofluorescence Spectrum Dynamics along the HRG-Induced Differentiation Pathway of MCF-7 Cells

    • Shin-ichi Morita,
    • Sota Takanezawa,
    • Michio Hiroshima,
    • Toshiyuki Mitsui,
    • Yukihiro Ozaki,
    • Yasushi Sako
    Cellular differentiation proceeds along complicated pathways, even when it is induced by extracellular signaling molecules. One of the major reasons for this complexity is the highly multidimensional internal dynamics of cells, which sometimes causes apparently stochastic responses in individual cells to extracellular stimuli. Therefore, to understand cell differentiation, it is necessary to monitor the internal dynamics of cells at single-cell resolution. Here, we used a Raman and autofluorescence spectrum analysis of single cells to detect dynamic changes in intracellular molecular components.
  • Article

    Polarized THG Microscopy Identifies Compositionally Different Lipid Droplets in Mammalian Cells

    • Godofredo Bautista,
    • Simon G. Pfisterer,
    • Mikko J. Huttunen,
    • Sanjeev Ranjan,
    • Kristiina Kanerva,
    • Elina Ikonen,
    • Martti Kauranen
    Cells store excess lipids as two major compounds, triacylglycerols (TAGs) and cholesteryl esters (CEs), inside lipid droplets (LDs). The degree of lipid ordering is considered to play a major role in the mobility and enzymatic processing of lipids in LDs. Here, we provide evidence that polarized third-harmonic generation (THG) microscopy distinguishes between native TAG- and CE-enriched LDs in cells due to the different ordering of the two lipid species. We first demonstrate that the responses from synthetic TAG- and CE-enriched LDs using THG microscopy with linear and circular polarizations differ according to their different intrinsic ordering.
  • Article

    A Comparative Mechanical Analysis of Plant and Animal Cells Reveals Convergence across Kingdoms

    • Pauline Durand-Smet,
    • Nicolas Chastrette,
    • Axel Guiroy,
    • Alain Richert,
    • Annick Berne-Dedieu,
    • Judit Szecsi,
    • Arezki Boudaoud,
    • Jean-Marie Frachisse,
    • Mohammed Bendahmane,
    • Oliver Hamant,
    • Atef Asnacios
    Plant and animals have evolved different strategies for their development. Whether this is linked to major differences in their cell mechanics remains unclear, mainly because measurements on plant and animal cells relied on independent experiments and setups, thus hindering any direct comparison. In this study we used the same micro-rheometer to compare animal and plant single cell rheology. We found that wall-less plant cells exhibit the same weak power law rheology as animal cells, with comparable values of elastic and loss moduli.
  • Article

    Molecular Determinants of Mechanical Properties of V. cholerae Biofilms at the Air-Liquid Interface

    • Emily C. Hollenbeck,
    • Jiunn C.N. Fong,
    • Ji Youn Lim,
    • Fitnat H. Yildiz,
    • Gerald G. Fuller,
    • Lynette Cegelski
    Biofilm formation increases both the survival and infectivity of Vibrio cholerae, the causative agent of cholera. V. cholerae is capable of forming biofilms on solid surfaces and at the air-liquid interface, termed pellicles. Known components of the extracellular matrix include the matrix proteins Bap1, RbmA, and RbmC, an exopolysaccharide termed Vibrio polysaccharide, and DNA. In this work, we examined a rugose strain of V. cholerae and its mutants unable to produce matrix proteins by interfacial rheology to compare the evolution of pellicle elasticity in real time to understand the molecular basis of matrix protein contributions to pellicle integrity and elasticity.
  • Article

    A Raman Microspectroscopy Study of Water and Trehalose in Spin-Dried Cells

    • Alireza Abazari,
    • Nilay Chakraborty,
    • Steven Hand,
    • Alptekin Aksan,
    • Mehmet Toner
    Long-term storage of desiccated nucleated mammalian cells at ambient temperature may be accomplished in a stable glassy state, which can be achieved by removal of water from the biological sample in the presence of glass-forming agents including trehalose. The stability of the glass may be compromised due to a nonuniform distribution of residual water and trehalose within and around the desiccated cells. Thus, quantification of water and trehalose contents at the single-cell level is critical for predicting the glass formation and stability for dry storage.

Channels and Transporters

  • Article

    Selecting Ions by Size in a Calcium Channel: The Ryanodine Receptor Case Study

    • Dirk Gillespie,
    • Le Xu,
    • Gerhard Meissner
    Many calcium channels can distinguish between ions of the same charge but different size. For example, when cations are in direct competition with each other, the ryanodine receptor (RyR) calcium channel preferentially conducts smaller cations such as Li+ and Na+ over larger ones such as K+ and Cs+. Here, we analyze the physical basis for this preference using a previously established model of RyR permeation and selectivity. Like other calcium channels, RyR has four aspartate residues in its GGGIGDE selectivity filter.


  • Article

    Area per Lipid and Cholesterol Interactions in Membranes from Separated Local-Field 13C NMR Spectroscopy

    • Avigdor Leftin,
    • Trivikram R. Molugu,
    • Constantin Job,
    • Klaus Beyer,
    • Michael F. Brown
    Investigations of lipid membranes using NMR spectroscopy generally require isotopic labeling, often precluding structural studies of complex lipid systems. Solid-state 13C magic-angle spinning NMR spectroscopy at natural isotopic abundance gives site-specific structural information that can aid in the characterization of complex biomembranes. Using the separated local-field experiment DROSS, we resolved 13C-1H residual dipolar couplings that were interpreted with a statistical mean-torque model.
  • Article

    Free Energy Landscape of Rim-Pore Expansion in Membrane Fusion

    • Herre Jelger Risselada,
    • Yuliya Smirnova,
    • Helmut Grubmüller
    The productive fusion pore in membrane fusion is generally thought to be toroidally shaped. Theoretical studies and recent experiments suggest that its formation, in some scenarios, may be preceded by an initial pore formed near the rim of the extended hemifusion diaphragm (HD), a rim-pore. This rim-pore is characterized by a nontoroidal shape that changes with size. To determine this shape as well as the free energy along the pathway of rim-pore expansion, we derived a simple analytical free energy model.
  • Article

    Transmembrane Protein Diffusion in Gel-Supported Dual-Leaflet Membranes

    • Chih-Ying Wang,
    • Reghan J. Hill
    Tools to measure transmembrane-protein diffusion in lipid bilayer membranes have advanced in recent decades, providing a need for predictive theoretical models that account for interleaflet leaflet friction on tracer mobility. Here we address the fully three-dimensional flows driven by a (nonprotruding) transmembrane protein embedded in a dual-leaflet membrane that is supported above and below by soft porous supports (e.g., hydrogel or extracellular matrix), each of which has a prescribed permeability and solvent viscosity.
  • Article

    Two Classes of Cholesterol Binding Sites for the β2AR Revealed by Thermostability and NMR

    • Deborah L. Gater,
    • Olivier Saurel,
    • Iordan Iordanov,
    • Wei Liu,
    • Vadim Cherezov,
    • Alain Milon
    Cholesterol binding to G protein-coupled receptors (GPCRs) and modulation of their activities in membranes is a fundamental issue for understanding their function. Despite the identification of cholesterol binding sites in high-resolution x-ray structures of the β2 adrenergic receptor (β2AR) and other GPCRs, the binding affinity of cholesterol for this receptor and exchange rates between the free and bound cholesterol remain unknown. In this study we report the existence of two classes of cholesterol binding sites in β2AR.
  • Article

    Neutron Reflectometry Studies Define Prion Protein N-terminal Peptide Membrane Binding

    • Anton P. Le Brun,
    • Cathryn L. Haigh,
    • Simon C. Drew,
    • Michael James,
    • Martin P. Boland,
    • Steven J. Collins
    The prion protein (PrP), widely recognized to misfold into the causative agent of the transmissible spongiform encephalopathies, has previously been shown to bind to lipid membranes with binding influenced by both membrane composition and pH. Aside from the misfolding events associated with prion pathogenesis, PrP can undergo various posttranslational modifications, including internal cleavage events. Alpha- and beta-cleavage of PrP produces two N-terminal fragments, N1 and N2, respectively, which interact specifically with negatively charged phospholipids at low pH.
  • Article

    Structural Insight into the Transmembrane Domain and the Juxtamembrane Region of the Erythropoietin Receptor in Micelles

    • Qingxin Li,
    • Ying Lei Wong,
    • Qiwei Huang,
    • CongBao Kang
    Erythropoietin receptor (EpoR) dimerization is an important step in erythrocyte formation. Its transmembrane domain (TMD) and juxtamembrane (JM) region are essential for signal transduction across the membrane. A construct compassing residues S212–P259 and containing the TMD and JM region of the human EpoR was purified and reconstituted in detergent micelles. The solution structure of the construct was determined in dodecylphosphocholine (DPC) micelles by solution NMR spectroscopy. Structural and dynamic studies demonstrated that the TMD and JM region are an α-helix in DPC micelles, whereas residues S212–D224 at the N-terminus of the construct are not structured.
  • Article

    How Cholesterol Could Be Drawn to the Cytoplasmic Leaf of the Plasma Membrane by Phosphatidylethanolamine

    • Ha Giang,
    • M. Schick
    In the mammalian plasma membrane, cholesterol can translocate rapidly between the exoplasmic and cytoplasmic leaves, so that its distribution between them should be given by the equality of its chemical potential in the leaves. Due to its favorable interaction with sphingomyelin, which is almost entirely in the outer leaf, one expects the great majority of cholesterol to be there also. Experimental results do not support this, implying that there is some mechanism attracting cholesterol to the inner leaf.

Molecular Machines, Motors and Nanoscale Biophysics

  • Article

    Identifying Transport Behavior of Single-Molecule Trajectories

    • Benjamin M. Regner,
    • Daniel M. Tartakovsky,
    • Terrence J. Sejnowski
    Models of biological diffusion-reaction systems require accurate classification of the underlying diffusive dynamics (e.g., Fickian, subdiffusive, or superdiffusive). We use a renormalization group operator to identify the anomalous (non-Fickian) diffusion behavior from a short trajectory of a single molecule. The method provides quantitative information about the underlying stochastic process, including its anomalous scaling exponent. The classification algorithm is first validated on simulated trajectories of known scaling.
  • Article

    Resonant Reflection Spectroscopy of Biomolecular Arrays in Muscle

    • Kevin W. Young,
    • Stojan Radic,
    • Evgeny Myslivets,
    • Shawn M. O’Connor,
    • Richard L. Lieber
    Sarcomeres, the functional units of contraction in striated muscle, are composed of an array of interdigitating protein filaments. Direct interaction between overlapping filaments generates muscular force, which produces animal movement. When filament length is known, sarcomere length successfully predicts potential force, even in whole muscles that contain billions of sarcomere units. Inability to perform in vivo sarcomere measurements with submicrometer resolution is a long-standing challenge in the muscle physiology field and has hampered studies of normal muscle function, adaptation, injury, aging, and disease, particularly in humans.
  • Article

    Fractal Globules: A New Approach to Artificial Molecular Machines

    • Vladik A. Avetisov,
    • Viktor A. Ivanov,
    • Dmitry A. Meshkov,
    • Sergei K. Nechaev
    The over-damped relaxation of elastic networks constructed by contact maps of hierarchically folded fractal (crumpled) polymer globules was investigated in detail. It was found that the relaxation dynamics of an anisotropic fractal globule is very similar to the behavior of biological molecular machines like motor proteins. When it is perturbed, the system quickly relaxes to a low-dimensional manifold, M, with a large basin of attraction and then slowly approaches equilibrium, not escaping M. Taking these properties into account, it is suggested that fractal globules, even those made by synthetic polymers, are artificial molecular machines that can transform perturbations into directed quasimechanical motion along a defined path.
  • Article

    The Dilated Cardiomyopathy-Causing Mutation ACTC E361G in Cardiac Muscle Myofibrils Specifically Abolishes Modulation of Ca2+ Regulation by Phosphorylation of Troponin I

    • Petr G. Vikhorev,
    • Weihua Song,
    • Ross Wilkinson,
    • O’Neal Copeland,
    • Andrew E. Messer,
    • Michael A. Ferenczi,
    • Steven B. Marston
    Phosphorylation of troponin I by protein kinase A (PKA) reduces Ca2+ sensitivity and increases the rate of Ca2+ release from troponin C and the rate of relaxation in cardiac muscle. In vitro experiments indicate that mutations that cause dilated cardiomyopathy (DCM) uncouple this modulation, but this has not been demonstrated in an intact contractile system. Using a Ca2+-jump protocol, we measured the effect of the DCM-causing mutation ACTC E361G on the equilibrium and kinetic parameters of Ca2+ regulation of contractility in single transgenic mouse heart myofibrils.
  • Article

    Smooth DNA Transport through a Narrowed Pore Geometry

    • Spencer Carson,
    • James Wilson,
    • Aleksei Aksimentiev,
    • Meni Wanunu
    Voltage-driven transport of double-stranded DNA through nanoscale pores holds much potential for applications in quantitative molecular biology and biotechnology, yet the microscopic details of translocation have proven to be challenging to decipher. Earlier experiments showed strong dependence of transport kinetics on pore size: fast regular transport in large pores (> 5 nm diameter), and slower yet heterogeneous transport time distributions in sub-5 nm pores, which imply a large positional uncertainty of the DNA in the pore as a function of the translocation time.

Proteins and Nucleic Acids

  • Article

    Electrostatic Channeling in P. falciparum DHFR-TS: Brownian Dynamics and Smoluchowski Modeling

    • Vincent T. Metzger,
    • Changsun Eun,
    • Peter M. Kekenes-Huskey,
    • Gary Huber,
    • J. Andrew McCammon
    We perform Brownian dynamics simulations and Smoluchowski continuum modeling of the bifunctional Plasmodium falciparum dihydrofolate reductase-thymidylate synthase (P. falciparum DHFR-TS) with the objective of understanding the electrostatic channeling of dihydrofolate generated at the TS active site to the DHFR active site. The results of Brownian dynamics simulations and Smoluchowski continuum modeling suggest that compared to Leishmania major DHFR-TS, P. falciparum DHFR-TS has a lower but significant electrostatic-mediated channeling efficiency (∼15–25%) at physiological pH (7.0) and ionic strength (150 mM).

Systems Biophysics

  • Article

    Stochastic Oscillations Induced by Intrinsic Fluctuations in a Self-Repressing Gene

    • Jingkui Wang,
    • Marc Lefranc,
    • Quentin Thommen
    Biochemical reaction networks are subjected to large fluctuations attributable to small molecule numbers, yet underlie reliable biological functions. Thus, it is important to understand how regularity can emerge from noise. Here, we study the stochastic dynamics of a self-repressing gene with arbitrarily long or short response time. We find that when the mRNA and protein half-lives are approximately equal to the gene response time, fluctuations can induce relatively regular oscillations in the protein concentration.
  • Article

    Juxtacrine Signaling Is Inherently Noisy

    • Tomer Yaron,
    • Yossi Cordova,
    • David Sprinzak
    Juxtacrine signaling is an important class of signaling systems that plays a crucial role in various developmental processes ranging from coordination of differentiation between neighboring cells to guiding axon growth during neurogenesis. Such signaling systems rely on the interaction between receptors on one cell and trans-membrane ligands on the membrane of a neighboring cell. Like other signaling systems, the ability of signal-receiving cells to accurately determine the concentration of ligands, is affected by stochastic diffusion processes.
  • Article

    The Robustness of Proofreading to Crowding-Induced Pseudo-Processivity in the MAPK Pathway

    • Thomas E. Ouldridge,
    • Pieter Rein ten Wolde
    Double phosphorylation of protein kinases is a common feature of signaling cascades. This motif may reduce cross-talk between signaling pathways because the second phosphorylation site allows for proofreading, especially when phosphorylation is distributive rather than processive. Recent studies suggest that phosphorylation can be pseudo-processive in the crowded cellular environment, since rebinding after the first phosphorylation is enhanced by slow diffusion. Here, we use a simple model with unsaturated reactants to show that specificity for one substrate over another drops as rebinding increases and pseudo-processive behavior becomes possible.
  • Article

    In Vivo Nonlinear Optical Imaging of Immune Responses: Tissue Injury and Infection

    • Yan Zeng,
    • Bo Yan,
    • Jin Xu,
    • Qiqi Sun,
    • Sicong He,
    • Jun Jiang,
    • Zilong Wen,
    • Jianan Y. Qu
    In this study, we demonstrate a noninvasive imaging approach based on multimodal nonlinear optical microscopy to in vivo image the responses of immune cells (neutrophils) to the tissue injury and bacterial infection in a zebrafish model. Specifically, the second harmonic generation from myosin thick filaments in sarcomere enabled a clear visualization of the muscle injury and infection. Two-photon excited fluorescence was used to track the behavior of the neutrophils that were transgenically labeled by red fluorescent protein.
  • Article

    Fibroblast Electrical Remodeling in Heart Failure and Potential Effects on Atrial Fibrillation

    • Martin Aguilar,
    • Xiao Yan Qi,
    • Hai Huang,
    • Philippe Comtois,
    • Stanley Nattel
    Fibroblasts are activated in heart failure (HF) and produce fibrosis, which plays a role in maintaining atrial fibrillation (AF). The effect of HF on fibroblast ion currents and its potential role in AF are unknown. Here, we used a patch-clamp technique to investigate the effects of HF on atrial fibroblast ion currents, and mathematical computation to assess the potential impact of this remodeling on atrial electrophysiology and arrhythmogenesis. Atrial fibroblasts were isolated from control and tachypacing-induced HF dogs.
  • Article

    Designing Cell-Targeted Therapeutic Proteins Reveals the Interplay between Domain Connectivity and Cell Binding

    • Avi Robinson-Mosher,
    • Jan-Hung Chen,
    • Jeffrey Way,
    • Pamela A. Silver
    The therapeutic efficacy of cytokines is often hampered by severe side effects due to their undesired binding to healthy cells. One strategy for overcoming this obstacle is to tether cytokines to antibodies or antibody fragments for targeted cell delivery. However, how to modulate the geometric configuration and relative binding affinity of the two domains for optimal activity remains an outstanding question. As a result, many antibody-cytokine complexes do not achieve the desired level of cell-targeted binding and activity.
  • Article

    Gap Junctions Suppress Electrical but Not [Ca2+] Heterogeneity in Resistance Arteries

    • Bjørn Olav Hald,
    • Donald G. Welsh,
    • Niels-Henrik Holstein-Rathlou,
    • Jens Chr. Brings Jacobsen
    Despite stochastic variation in the molecular composition and morphology of individual smooth muscle and endothelial cells, the membrane potential along intact microvessels is remarkably uniform. This is crucial for coordinated vasomotor responses. To investigate how this electrical homogeneity arises, a virtual arteriole was developed that introduces variation in the activities of ion-transport proteins between cells. By varying the level of heterogeneity and subpopulations of gap junctions (GJs), the resulting simulations shows that GJs suppress electrical variation but can only reduce cytosolic [Ca2+] variation.


  • Correction


    2014. I. Budin, N. Prywes, N. Zhang, J. W. Szostak. Chain-Length Heterogeneity Allows for the Assembly of Fatty Acid Vesicles in Dilute Solutions. Biophys. J. 107:1582–1590.
  • Correction


    2014. C. A. Villalba-Galea. Hv1 Proton Channel Opening Is Preceded by a Voltage-Independent Transition. Biophys J. 107(7):1564–1572