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Mark R Worden - One of the best experts on this subject based on the ideXlab platform.
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voltage dependent closure of porb class ii porin from neisseria meningitidis investigated using impedance spectroscopy in a tethered bilayer Lipid Membrane interface
Journal of Colloid and Interface Science, 2013Co-Authors: Sachin R Jadhav, Yi Zheng, Michael R Garavito, Kota Sreenivasa Rao, Mark R WordenAbstract:Abstract Electrochemical impedance spectroscopy (EIS) was used to characterize voltage-dependent closure of PorB class II (PorBII) porin from Neisseria meningitidis incorporated in a tethered bilayer Lipid Membrane (tBLM). The tBLM’s lower leaflet was fabricated by depositing a self assembled monolayer (SAM) of 1,2-dipalmitoyl- sn -glycero-3-phosphothioethanol (DPPTE) on a gold electrode, and the upper leaflet was formed by depositing1,2-dioleoyl- sn -glycero-3-phoshocholine (DOPC) liposomes. At 0 mV bias DC potential, incorporation of PorBII decreased the Membrane resistance ( R m ) from 2.5 MΩ cm 2 to 0.6 MΩ cm 2 , giving a Δ R m of 1.9 MΩ cm 2 and a normalized Δ R m (Δ R m divided by the R m of the tBLM without PorBII) of 76%. When the bias DC potential was increased to 200 mV, the normalized Δ R m value decreased to 20%. The effect of applied voltage on Δ R m was completely reversible, suggesting voltage-dependent closure of PorBII. The voltage dependence of PorBII was further studied in a planar bilayer Lipid Membrane made from 1,2-diphytanoyl- sn -glycero-3-phosphocholine (DPhytPC). Following a single insertion event, PorBII exhibited multiple conductance states, with reversible, voltage-dependent closure of PorBII porin occurring at high transMembrane potentials. The trimetric porin closed in three discrete steps, each step corresponding to closure of one conducting monomer unit. The most probable single channel conductance was 4.2 nS. The agreement between results obtained with the tBLM and pBLM platforms demonstrates the utility of EIS to screen channel proteins immobilized in tBLM for voltage-gated behavior.
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functional characterization of porb class ii porin from neisseria meningitidis using a tethered bilayer Lipid Membrane
Biosensors and Bioelectronics, 2008Co-Authors: Sachin R Jadhav, Yi Zheng, Michael R Garavito, Mark R WordenAbstract:PorB class II from Neisseria meningitidis is a pore-forming, outer-Membrane protein that can translocate to the host-cell Membrane during Neisserial infections. This report describes development of tethered bilayer Lipid Membrane (tBLM) system to measure PorB conductance properties. The tBLM was fabricated by depositing a self-assembled monolayer of 1,2-dipalmitoyl-sn-glycero-3-phosphothioethanol (DPPTE) tethering Lipid on a gold electrode and then using 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) liposomes to deposit the upper tBLM leaflet. Electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) were used to monitor tBLM formation and subsequent PorB incorporation. The highly insulating tBLM exhibited a Membrane resistance and capacitance of 2.5MOmegacm(2) and 0.7 microF/cm(2), respectively. PorB was incorporated into the tBLM in an active conformation, as evidenced by its mediation of ion passage and the decrease in Membrane impedance. After PorB incorporation, the Membrane resistance decreased to 0.6MOmegacm(2). As expected, the PorB channel was found to be non-selective, allowing the transport of both cations and anions. Cyclic voltammetry indicated that ferricyanide ions can also pass through the pores. The PorB-containing biomimetic interface developed in this study could potentially be used to screen for compounds that modulate PorB activity.
Erkang Wang - One of the best experts on this subject based on the ideXlab platform.
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an electrochemical study on the interaction of surfactin with a supported bilayer Lipid Membrane on a glassy carbon electrode
Journal of Electroanalytical Chemistry, 2005Co-Authors: Xiaohua Liu, Weimin Huang, Erkang WangAbstract:Surfactin is an acidic lipopeptide with amphiphilic character, which can interact with a bioMembrane. The interaction of surfactin with a supported bilayer Lipid Membrane on a glassy carbon electrode (GCE) was investigated by cyclic voltammetry and ac impedance spectroscopy in this paper. Surfactin could induce pores in the bilayer Lipid Membrane and even cause the destruction of the Membrane. The mechanism of the interaction of surfactin with the supported bilayer Lipid Membrane was studied. The insertion of surfactin into the Lipid Membrane was the first step in the formation of pores. Subsequently, surfactin aggregated in the Lipid Membrane to form the pores. Along with the transition of surfactin molecules from the outer to the inner layer, which was connected to the surface of the GCE, the Lipid Membrane was destroyed.
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hydrogen peroxide biosensor based on microperoxidase 11 entrapped in Lipid Membrane
Biosensors and Bioelectronics, 2003Co-Authors: Weimin Huang, Zheling Zhang, Xiaojun Han, Jianguo Wang, Jilin Tang, Shaojun Dong, Jianbo Jia, Erkang WangAbstract:A highly catalytic activity microperoxidase-11 (MP-11) biosensor for H(2)O(2) was developed to immobilizing the heme peptide in didodecyldimethylammonium bromide (DDAB) Lipid Membrane. The enzyme electrode thus obtained responded to H(2)O(2) without electron mediator or promoter, at a potential of +0.10 V versus Agmid R:AgCl. A linear calibration curve is obtained over the range from 2.0 x 10(-5) to 2.4 x 10(-3) M. The biosensor responds to hydrogen peroxide in 15 s and has a detection limit of 8 x 10(-7) M (S/N=3) Providing a natural environment with Lipid Membrane for protein immobilization and maintenance of protein functions is a suitable option for the design of biosensors.
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Concentration-dependent behavior of nisin interaction with supported bilayer Lipid Membrane.
Biophysical chemistry, 2002Co-Authors: Weimin Huang, Zheling Zhang, Xiaojun Han, Jianguo Wang, Jilin Tang, Shaojun Dong, Erkang WangAbstract:Nisin is a positively charged antibacterial peptide that binds to the negatively charged Membranes of gram-positive bacteria. The initial interaction of the peptide with the model Membrane of negatively charged DPPG (dipalmitoylphosphatidylglycerol) was studied by cyclic voltammetry and a.c. impedance spectroscopy. Nisin could induce pores in the supported bilayer Lipid Membrane, thus, it led to the marker ions Fe(CN)(6)(3-/4-) crossing the Lipid Membrane and giving the redox reaction on the glassy carbon electrode (GCE). Experimental results suggested that the pore formation on supported bilayer Lipid Membrane was dependent on the concentration of nisin and it included three main concentration stages: low, middling, high concentration.
Sachin R Jadhav - One of the best experts on this subject based on the ideXlab platform.
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voltage dependent closure of porb class ii porin from neisseria meningitidis investigated using impedance spectroscopy in a tethered bilayer Lipid Membrane interface
Journal of Colloid and Interface Science, 2013Co-Authors: Sachin R Jadhav, Yi Zheng, Michael R Garavito, Kota Sreenivasa Rao, Mark R WordenAbstract:Abstract Electrochemical impedance spectroscopy (EIS) was used to characterize voltage-dependent closure of PorB class II (PorBII) porin from Neisseria meningitidis incorporated in a tethered bilayer Lipid Membrane (tBLM). The tBLM’s lower leaflet was fabricated by depositing a self assembled monolayer (SAM) of 1,2-dipalmitoyl- sn -glycero-3-phosphothioethanol (DPPTE) on a gold electrode, and the upper leaflet was formed by depositing1,2-dioleoyl- sn -glycero-3-phoshocholine (DOPC) liposomes. At 0 mV bias DC potential, incorporation of PorBII decreased the Membrane resistance ( R m ) from 2.5 MΩ cm 2 to 0.6 MΩ cm 2 , giving a Δ R m of 1.9 MΩ cm 2 and a normalized Δ R m (Δ R m divided by the R m of the tBLM without PorBII) of 76%. When the bias DC potential was increased to 200 mV, the normalized Δ R m value decreased to 20%. The effect of applied voltage on Δ R m was completely reversible, suggesting voltage-dependent closure of PorBII. The voltage dependence of PorBII was further studied in a planar bilayer Lipid Membrane made from 1,2-diphytanoyl- sn -glycero-3-phosphocholine (DPhytPC). Following a single insertion event, PorBII exhibited multiple conductance states, with reversible, voltage-dependent closure of PorBII porin occurring at high transMembrane potentials. The trimetric porin closed in three discrete steps, each step corresponding to closure of one conducting monomer unit. The most probable single channel conductance was 4.2 nS. The agreement between results obtained with the tBLM and pBLM platforms demonstrates the utility of EIS to screen channel proteins immobilized in tBLM for voltage-gated behavior.
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functional characterization of porb class ii porin from neisseria meningitidis using a tethered bilayer Lipid Membrane
Biosensors and Bioelectronics, 2008Co-Authors: Sachin R Jadhav, Yi Zheng, Michael R Garavito, Mark R WordenAbstract:PorB class II from Neisseria meningitidis is a pore-forming, outer-Membrane protein that can translocate to the host-cell Membrane during Neisserial infections. This report describes development of tethered bilayer Lipid Membrane (tBLM) system to measure PorB conductance properties. The tBLM was fabricated by depositing a self-assembled monolayer of 1,2-dipalmitoyl-sn-glycero-3-phosphothioethanol (DPPTE) tethering Lipid on a gold electrode and then using 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) liposomes to deposit the upper tBLM leaflet. Electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) were used to monitor tBLM formation and subsequent PorB incorporation. The highly insulating tBLM exhibited a Membrane resistance and capacitance of 2.5MOmegacm(2) and 0.7 microF/cm(2), respectively. PorB was incorporated into the tBLM in an active conformation, as evidenced by its mediation of ion passage and the decrease in Membrane impedance. After PorB incorporation, the Membrane resistance decreased to 0.6MOmegacm(2). As expected, the PorB channel was found to be non-selective, allowing the transport of both cations and anions. Cyclic voltammetry indicated that ferricyanide ions can also pass through the pores. The PorB-containing biomimetic interface developed in this study could potentially be used to screen for compounds that modulate PorB activity.
Roderick Mackinnon - One of the best experts on this subject based on the ideXlab platform.
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mechanosensitivity is mediated directly by the Lipid Membrane in traak and trek1 k channels
Proceedings of the National Academy of Sciences of the United States of America, 2014Co-Authors: Stephen G Brohawn, Zhenwei Su, Roderick MackinnonAbstract:Mechanosensitive ion channels underlie neuronal responses to physical forces in the sensation of touch, hearing, and other mechanical stimuli. The fundamental basis of force transduction in eukaryotic mechanosensitive ion channels is unknown. Are mechanical forces transmitted directly from Membrane to channel as in prokaryotic mechanosensors or are they mediated through macromolecular tethers attached to the channel? Here we show in cells that the K+ channel TRAAK (K2P4.1) is responsive to mechanical forces similar to the ion channel Piezo1 and that mechanical activation of TRAAK can electrically counter Piezo1 activation. We then show that the biophysical origins of force transduction in TRAAK and TREK1 (K2P2.1) two-pore domain K+ (K2P) channels come from the Lipid Membrane, not from attached tethers. These findings extend the “force-from-Lipid” principle established for prokaryotic mechanosensitive channels MscL and MscS to these eukaryotic mechanosensitive K+ channels.
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voltage dependent k channel gating and voltage sensor toxin sensitivity depend on the mechanical state of the Lipid Membrane
Proceedings of the National Academy of Sciences of the United States of America, 2008Co-Authors: Daniel F Schmidt, Roderick MackinnonAbstract:Voltage-dependent K+ (Kv) channels underlie action potentials through gating conformational changes that are driven by Membrane voltage. In this study of the paddle chimera Kv channel, we demonstrate that the rate of channel opening, the voltage dependence of the open probability, and the maximum achievable open probability depend on the Lipid Membrane environment. The activity of the voltage sensor toxin VsTx1, which interferes with voltage-dependent gating by partitioning into the Membrane and binding to the channel, also depends on the Membrane. Membrane environmental factors that influence channel function are divisible into two general categories: Lipid compositional and mechanical state. The mechanical state can have a surprisingly large effect on the function of a voltage-dependent K+ channel, including its pharmacological interaction with voltage sensor toxins. The dependence of VSTx1 activity on the mechanical state of the Membrane leads us to hypothesize that voltage sensor toxins exert their effect by perturbing the interaction forces that exist between the channel and the Membrane.
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Structure of the KvAP voltage-dependent K+ channel and its dependence on the Lipid Membrane
Proceedings of the National Academy of Sciences, 2005Co-Authors: S. Y. Lee, A. Lee, Roderick MackinnonAbstract:Voltage-dependent ion channels gate open in response to changes in cell Membrane voltage. This form of gating permits the propagation of action potentials. We present two structures of the voltage-dependent K(+) channel KvAP, in complex with monoclonal Fv fragments (3.9 A) and without antibody fragments (8 A). We also studied KvAP with disulfide cross-bridges in Lipid Membranes. Analyzing these data in the context of the crystal structure of Kv1.2 and EPR data on KvAP we reach the following conclusions: (i) KvAP is similar in structure to Kv1.2 with a very modest difference in the orientation of its voltage sensor; (ii) mAb fragments are not the source of non-native conformations of KvAP in crystal structures; (iii) because KvAP contains separate loosely adherent domains, a Lipid Membrane is required to maintain their correct relative orientations, and (iv) the model of KvAP is consistent with the proposal of voltage sensing through the movement of an arginine-containing helix-turn-helix element at the protein-Lipid interface.
Weimin Huang - One of the best experts on this subject based on the ideXlab platform.
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an electrochemical study on the interaction of surfactin with a supported bilayer Lipid Membrane on a glassy carbon electrode
Journal of Electroanalytical Chemistry, 2005Co-Authors: Xiaohua Liu, Weimin Huang, Erkang WangAbstract:Surfactin is an acidic lipopeptide with amphiphilic character, which can interact with a bioMembrane. The interaction of surfactin with a supported bilayer Lipid Membrane on a glassy carbon electrode (GCE) was investigated by cyclic voltammetry and ac impedance spectroscopy in this paper. Surfactin could induce pores in the bilayer Lipid Membrane and even cause the destruction of the Membrane. The mechanism of the interaction of surfactin with the supported bilayer Lipid Membrane was studied. The insertion of surfactin into the Lipid Membrane was the first step in the formation of pores. Subsequently, surfactin aggregated in the Lipid Membrane to form the pores. Along with the transition of surfactin molecules from the outer to the inner layer, which was connected to the surface of the GCE, the Lipid Membrane was destroyed.
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hydrogen peroxide biosensor based on microperoxidase 11 entrapped in Lipid Membrane
Biosensors and Bioelectronics, 2003Co-Authors: Weimin Huang, Zheling Zhang, Xiaojun Han, Jianguo Wang, Jilin Tang, Shaojun Dong, Jianbo Jia, Erkang WangAbstract:A highly catalytic activity microperoxidase-11 (MP-11) biosensor for H(2)O(2) was developed to immobilizing the heme peptide in didodecyldimethylammonium bromide (DDAB) Lipid Membrane. The enzyme electrode thus obtained responded to H(2)O(2) without electron mediator or promoter, at a potential of +0.10 V versus Agmid R:AgCl. A linear calibration curve is obtained over the range from 2.0 x 10(-5) to 2.4 x 10(-3) M. The biosensor responds to hydrogen peroxide in 15 s and has a detection limit of 8 x 10(-7) M (S/N=3) Providing a natural environment with Lipid Membrane for protein immobilization and maintenance of protein functions is a suitable option for the design of biosensors.
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Concentration-dependent behavior of nisin interaction with supported bilayer Lipid Membrane.
Biophysical chemistry, 2002Co-Authors: Weimin Huang, Zheling Zhang, Xiaojun Han, Jianguo Wang, Jilin Tang, Shaojun Dong, Erkang WangAbstract:Nisin is a positively charged antibacterial peptide that binds to the negatively charged Membranes of gram-positive bacteria. The initial interaction of the peptide with the model Membrane of negatively charged DPPG (dipalmitoylphosphatidylglycerol) was studied by cyclic voltammetry and a.c. impedance spectroscopy. Nisin could induce pores in the supported bilayer Lipid Membrane, thus, it led to the marker ions Fe(CN)(6)(3-/4-) crossing the Lipid Membrane and giving the redox reaction on the glassy carbon electrode (GCE). Experimental results suggested that the pore formation on supported bilayer Lipid Membrane was dependent on the concentration of nisin and it included three main concentration stages: low, middling, high concentration.