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Zhan Chen - One of the best experts on this subject based on the ideXlab platform.
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Sum Frequency Generation of Interfacial Lipid Monolayers Shows Polarization Dependence on Experimental Geometries
Langmuir : the ACS journal of surfaces and colloids, 2016Co-Authors: Yong Hao, Xiaofeng Han, Zhirui Guo, Zhan ChenAbstract:Sum frequency generation (SFG) vibrational spectroscopy has been widely employed to investigate molecular structures of biological surfaces and interfaces including Model Cell membranes. A variety of lipid monolayers or bilayers serving as Model Cell membranes and their interactions with many different molecules have been extensively studied using SFG. Here, we conducted an in-depth investigation on polarization-dependent SFG signals collected from interfacial lipid monolayers using different experimental geometries, i.e., the prism geometry (total internal reflection) and the window geometry (external reflection). The different SFG spectral features of interfacial lipid monolayers detected using different experimental geometries are due to the interplay between the varied Fresnel coefficients and second-order nonlinear susceptibility tensor terms of different vibrational modes (i.e., ss and as modes of methyl groups), which were analyzed in detail in this study. Therefore, understanding the interplay bet...
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Observing a Model Ion Channel Gating Action in Model Cell Membranes in Real Time in Situ: Membrane Potential Change Induced Alamethicin Orientation Change
2016Co-Authors: Feng Wei, Pei Yang, Joshua Jasensky, Andrew P. Boughton, Zhan ChenAbstract:Ion channels play crucial roles in transport and regulatory functions of living Cells. Understanding the gating mechanisms of these channels is important to understanding and treating diseases that have been linked to ion channels. One potential Model peptide for studying the mechanism of ion channel gating is alamethicin, which adopts a split α/310-helix structure and responds to changes in electric potential. In this study, sum frequency generation vibrational spectroscopy (SFG-VS), supplemented by attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR), has been applied to characterize interactions between alamethicin (a Model for larger channel proteins) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) lipid bilayers in the presence of an electric potential across the membrane. The membrane potential difference was controlled by changing the pH of the solution in contact with the bilayer and was measured using fluorescence spectroscopy. The orientation angle of alamethicin in POPC lipid bilayers was then determined at different pH values using polarized SFG amide I spectra. Assuming that all molecules adopt the same orientation (a δ distribution), at pH = 6.7 the α-helix at the N-terminus and the 310-helix at the C-terminus tilt at about 72° (θ1) and 50° (θ2) versus the surface normal, respectively. When pH increases to 11.9, θ1 and θ2 decrease to 56.5° and 45°, respectively. The δ distribution assumption was verified using a combination of SFG and ATR-FTIR measurements, which showed a quite narrow distribution in the angle of θ1 for both pH conditions. This indicates that all alamethicin molecules at the surface adopt a nearly identical orientation in POPC lipid bilayers. The localized pH change in proximity to the bilayer modulates the membrane potential and thus induces a decrease in both the tilt and the bend angles of the two helices in alamethicin. This is the first reported application of SFG to the study of Model ion channel gating mechanisms in Model Cell membranes
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Interactions of alamethicin with Model Cell membranes investigated using sum frequency generation vibrational spectroscopy
2015Co-Authors: Khoi Tan Nguyen, Zhan ChenAbstract:Structures of membrane associated peptides and molecular interactions between peptides and Cell membrane bilayers govern biological functions of these peptides. Sum frequency generation (SFG) vibrational spectroscopy has been demonstrated to be a powerful technique to study such structures and interactions at the molecular level. In this research, SFG has been applied, supplemented by attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR), to characterize interactions between alamethicin (a Model for larger channel proteins) and different lipid bilayers in the absence of membrane potential. The orientation of alamethicin in lipid bilayers has been determined using SFG amide I spectra detected with different polarization combinations. It was found that alamethicin adopts a mixed α-helical and 310- helical structure in fluid-phase lipid bilayers. The helix (mainly α-helix) at the N-terminus tilts at about 63 ° versus the surface normal in a fluid-phase 1,2-Dimyristoyl-D54-sn-Glycero-3-Phosphocholine-1,1,2,2-D4-N,N,N-trimethyl-D9 (d-DMPC)/ 1,2-Dimyristoyl-sn-Glycero-3-Phosphocholine (DMPC) bilayer. The 310 helix at the C-terminus (beyond the Pro14 residue) tilts at about 43 ° versus the surface normal. This is the first time to apply SFG to study a 310 helix experimentally. When interacting with a gel-phase lipid bilayer, alamethicin lies down on the gel-phase bilayer surface and/or aggregates, which does not have significant insertion into the lipid bilayer. 1
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interfacial fresnel coefficients and molecular structures of Model Cell membranes from a lipid monolayer to a lipid bilayer
Journal of Physical Chemistry C, 2014Co-Authors: Xiaofeng Han, John N Myers, Zhan ChenAbstract:During the Model membrane formation process from a lipid monolayer of 1,2-dipalmitoyl-sn-glycero-3-phospho-(1′-rac-glycerol) (sodium salt) (DPPG) in air to a lipid bilayer of DPPG/deuterated-DPPG (dDPPG) on water, the intensity of sum frequency generation (SFG) vibrational signals from the DPPG molecules increased by ∼34 times. The increased signal intensity could be caused by inherently different molecular ordering of lipid molecules in the monolayer/bilayer or by optical effects induced by different contacting mediums (prism in air and prism contacting water) (see Figure 2). We resolve the two possibilities by analyzing tilt angles of the methyl groups at DPPG hydrophobic ends for the monolayer/bilayer which reflect the molecular ordering information and by evaluating the Fresnel coefficients which reflect the contacting medium-induced optical effect. DPPG molecules were more ordered after transformation from a lipid monolayer in air to a lipid bilayer on water, which induced a slight signal increase. T...
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Interaction of Polyethylenimine with Model Cell Membranes Studied by Linear and Nonlinear Spectroscopic Techniques
2014Co-Authors: Chi Zhang, Zhan ChenAbstract:Polyethylenimine (PEI) has been widely used as a transfection agent for gene delivery, but it is cytotoxic and can lead to Cell apoptosis. Although several apoptotic mechanisms have been proposed, a molecular level understanding of PEI/Cell membrane interaction can help develop further insight into such cytotoxicity. We combined sum frequency generation (SFG) vibrational spectroscopy and attenuated total-internal reflection Fourier transform infrared (ATR-FTIR) spectroscopy to study the effect of PEI on lipid transbilayer movement in supported bilayers (serving as Model Cell membranes) as a function of lipid composition, PEI concentration, and temperature. For both dipalmitoylphosphatidylglycerol (DPPG) and distearoylphosphatidylcholine (DSPC) bilayers, PEI molecules showed no significant effect on lipid translocation at room temperature (21 °C). In contrast, significant lipid translocation was observed near the physiological temperature (39 °C), indicating the ability of PEI to induce lipid translocation in both negatively charged and zwitterionic lipid bilayers, without the assistance of membrane proteins. Furthermore, results showed that PEI had strong interactions with negatively charged DPPG and weak interactions with zwitterionic DSPC. Concentration-dependent studies indicated that the lipid translocation rate had a linear dependence on the PEI concentration in the subphase. The effects of branched and linear PEIs were compared in the study, showing that branched PEI had a greater effect on the lipid translocation rate due to the higher charge density, which might be a possible indication of higher toxicity. ATR-FTIR spectroscopy verified that the results observed in SFG were mainly caused by lipid translocation, not bilayer damage or removal from the substrate. The combined SFG and ATR-FTIR study provides a powerful method to examine molecular interactions between lipid bilayers and polyelectrolytes at a molecular level. The results can help to develop further understanding on PEI’s cytotoxicity in biological systems
Tetsuya Haruyama - One of the best experts on this subject based on the ideXlab platform.
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post synapse Model Cell for synaptic glutamate receptor glur based biosensing strategy and engineering to maximize ligand gated ion flux achieving high signal to noise ratio
Sensors, 2012Co-Authors: Akito Tateishi, Satoshi Migita, Kari Keinanen, Sarah K Coleman, Tetsuya HaruyamaAbstract:Cell-based biosensing is a “smart” way to obtain efficacy-information on the effect of applied chemical on Cellular biological cascade. We have proposed an engineered post-synapse Model Cell-based biosensors to investigate the effects of chemicals on ionotropic glutamate receptor (GluR), which is a focus of attention as a molecular target for clinical neural drug discovery. The engineered Model Cell has several advantages over native Cells, including improved ease of handling and better reproducibility in the application of Cell-based biosensors. However, in general, Cell-based biosensors often have low signal-to-noise (S/N) ratios due to the low level of Cellular responses. In order to obtain a higher S/N ratio in Model Cells, we have attempted to design a tactic Model Cell with elevated Cellular response. We have revealed that the increase GluR expression level is not directly connected to the amplification of Cellular responses because the saturation of surface expression of GluR, leading to a limit on the total ion influx. Furthermore, coexpression of GluR with a voltage-gated potassium channel increased Ca2+ ion influx beyond levels obtained with saturating amounts of GluR alone. The construction of Model Cells based on strategy of amplifying ion flux per individual receptors can be used to perform smart Cell-based biosensing with an improved S/N ratio.
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discerning data analysis methods to clarify agonistic antagonistic actions on the ion flux assay of ligand gated ionotropic glutamate receptor on engineered post synapse Model Cells
Journal of Biosensors and Bioelectronics, 2011Co-Authors: Akito Tateishi, Satoshi Migita, Kari Keinanen, Michael Cauchi, Chisato Tanoue, Sarah K Coleman, Shinya Ikeno, Conrad Bessant, Tetsuya HaruyamaAbstract:Cell-based experiments provide the efficacy of chemicals through the biological function. The authors have described post-synapse Model Cell-based assay based on qualified analysis for neural drug discoveries. However, in general, Cell-based assays often include data fluctuation. This may be a barrier preventing the performance for various practical purposes. In this study, we tried discerning data analysis for clarify the chemical action to the ionotoropic glutamate receptor (GluR), whereby an ion-flux assay of post-synapse Model Cells is performed and are analyzed based on a chemometrics approach. The dynamic behavior of the GluR of post-synapse Model Cell was assayed with multivariate data analysis methods namely hierarchical cluster analysis (HCA) and principal component analysis (PCA). By using HCA, we can identify and remove the non-responding samples. By using PCA, the effect of chemicals on the dynamic behavior of ion flux through GluR can be recognized clearly; as either agonist or antagonist. As shown in the results, the GluR-based assay by post-synapse Model Cell with data analysis methods provide a sodium influx profile which is derived by an agonists or antagonists application. By employing the data analysis methods, PCA and HCA, it is possible to develop a smart Cellular biosensing system for qualified analysis.
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engineered synapse Model Cell genetic construction and chemical evaluation for reproducible high throughput analysis
Analytical and Bioanalytical Chemistry, 2010Co-Authors: Satoshi Migita, Akito Tateishi, Kari Keinanen, Tetsuya HaruyamaAbstract:Bioassay Models of neural functions must lend themselves to high-throughput analysis in neural drug discovery. However, smart analysis methods for these functions have not yet been fully established. Here, we describe the development of a synapse Model for Cell-based biosensing. The engineered synapse Model Cell expresses ionotropic glutamate receptor on its surface, like the neural postsynaptic membrane. The advantages of the Model Cell are the ease of handling and reproducibility as compared with the cultured neural Cell, and it can be employed to evaluate receptor function through ion flux analysis. The agonist-induced sodium influx was monitored as an agonist concentration-dependent increase in the observed fluorescence signal. Furthermore, we found that our Model Cell enables the correction of uneven Cellular signal levels using a reporter system. Our engineered synapse Model Cell can be employed as a powerful tool for the screening of lead substances in pharmaceutical high-throughput analysis.
Satoshi Migita - One of the best experts on this subject based on the ideXlab platform.
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post synapse Model Cell for synaptic glutamate receptor glur based biosensing strategy and engineering to maximize ligand gated ion flux achieving high signal to noise ratio
Sensors, 2012Co-Authors: Akito Tateishi, Satoshi Migita, Kari Keinanen, Sarah K Coleman, Tetsuya HaruyamaAbstract:Cell-based biosensing is a “smart” way to obtain efficacy-information on the effect of applied chemical on Cellular biological cascade. We have proposed an engineered post-synapse Model Cell-based biosensors to investigate the effects of chemicals on ionotropic glutamate receptor (GluR), which is a focus of attention as a molecular target for clinical neural drug discovery. The engineered Model Cell has several advantages over native Cells, including improved ease of handling and better reproducibility in the application of Cell-based biosensors. However, in general, Cell-based biosensors often have low signal-to-noise (S/N) ratios due to the low level of Cellular responses. In order to obtain a higher S/N ratio in Model Cells, we have attempted to design a tactic Model Cell with elevated Cellular response. We have revealed that the increase GluR expression level is not directly connected to the amplification of Cellular responses because the saturation of surface expression of GluR, leading to a limit on the total ion influx. Furthermore, coexpression of GluR with a voltage-gated potassium channel increased Ca2+ ion influx beyond levels obtained with saturating amounts of GluR alone. The construction of Model Cells based on strategy of amplifying ion flux per individual receptors can be used to perform smart Cell-based biosensing with an improved S/N ratio.
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discerning data analysis methods to clarify agonistic antagonistic actions on the ion flux assay of ligand gated ionotropic glutamate receptor on engineered post synapse Model Cells
Journal of Biosensors and Bioelectronics, 2011Co-Authors: Akito Tateishi, Satoshi Migita, Kari Keinanen, Michael Cauchi, Chisato Tanoue, Sarah K Coleman, Shinya Ikeno, Conrad Bessant, Tetsuya HaruyamaAbstract:Cell-based experiments provide the efficacy of chemicals through the biological function. The authors have described post-synapse Model Cell-based assay based on qualified analysis for neural drug discoveries. However, in general, Cell-based assays often include data fluctuation. This may be a barrier preventing the performance for various practical purposes. In this study, we tried discerning data analysis for clarify the chemical action to the ionotoropic glutamate receptor (GluR), whereby an ion-flux assay of post-synapse Model Cells is performed and are analyzed based on a chemometrics approach. The dynamic behavior of the GluR of post-synapse Model Cell was assayed with multivariate data analysis methods namely hierarchical cluster analysis (HCA) and principal component analysis (PCA). By using HCA, we can identify and remove the non-responding samples. By using PCA, the effect of chemicals on the dynamic behavior of ion flux through GluR can be recognized clearly; as either agonist or antagonist. As shown in the results, the GluR-based assay by post-synapse Model Cell with data analysis methods provide a sodium influx profile which is derived by an agonists or antagonists application. By employing the data analysis methods, PCA and HCA, it is possible to develop a smart Cellular biosensing system for qualified analysis.
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engineered synapse Model Cell genetic construction and chemical evaluation for reproducible high throughput analysis
Analytical and Bioanalytical Chemistry, 2010Co-Authors: Satoshi Migita, Akito Tateishi, Kari Keinanen, Tetsuya HaruyamaAbstract:Bioassay Models of neural functions must lend themselves to high-throughput analysis in neural drug discovery. However, smart analysis methods for these functions have not yet been fully established. Here, we describe the development of a synapse Model for Cell-based biosensing. The engineered synapse Model Cell expresses ionotropic glutamate receptor on its surface, like the neural postsynaptic membrane. The advantages of the Model Cell are the ease of handling and reproducibility as compared with the cultured neural Cell, and it can be employed to evaluate receptor function through ion flux analysis. The agonist-induced sodium influx was monitored as an agonist concentration-dependent increase in the observed fluorescence signal. Furthermore, we found that our Model Cell enables the correction of uneven Cellular signal levels using a reporter system. Our engineered synapse Model Cell can be employed as a powerful tool for the screening of lead substances in pharmaceutical high-throughput analysis.
Akito Tateishi - One of the best experts on this subject based on the ideXlab platform.
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post synapse Model Cell for synaptic glutamate receptor glur based biosensing strategy and engineering to maximize ligand gated ion flux achieving high signal to noise ratio
Sensors, 2012Co-Authors: Akito Tateishi, Satoshi Migita, Kari Keinanen, Sarah K Coleman, Tetsuya HaruyamaAbstract:Cell-based biosensing is a “smart” way to obtain efficacy-information on the effect of applied chemical on Cellular biological cascade. We have proposed an engineered post-synapse Model Cell-based biosensors to investigate the effects of chemicals on ionotropic glutamate receptor (GluR), which is a focus of attention as a molecular target for clinical neural drug discovery. The engineered Model Cell has several advantages over native Cells, including improved ease of handling and better reproducibility in the application of Cell-based biosensors. However, in general, Cell-based biosensors often have low signal-to-noise (S/N) ratios due to the low level of Cellular responses. In order to obtain a higher S/N ratio in Model Cells, we have attempted to design a tactic Model Cell with elevated Cellular response. We have revealed that the increase GluR expression level is not directly connected to the amplification of Cellular responses because the saturation of surface expression of GluR, leading to a limit on the total ion influx. Furthermore, coexpression of GluR with a voltage-gated potassium channel increased Ca2+ ion influx beyond levels obtained with saturating amounts of GluR alone. The construction of Model Cells based on strategy of amplifying ion flux per individual receptors can be used to perform smart Cell-based biosensing with an improved S/N ratio.
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discerning data analysis methods to clarify agonistic antagonistic actions on the ion flux assay of ligand gated ionotropic glutamate receptor on engineered post synapse Model Cells
Journal of Biosensors and Bioelectronics, 2011Co-Authors: Akito Tateishi, Satoshi Migita, Kari Keinanen, Michael Cauchi, Chisato Tanoue, Sarah K Coleman, Shinya Ikeno, Conrad Bessant, Tetsuya HaruyamaAbstract:Cell-based experiments provide the efficacy of chemicals through the biological function. The authors have described post-synapse Model Cell-based assay based on qualified analysis for neural drug discoveries. However, in general, Cell-based assays often include data fluctuation. This may be a barrier preventing the performance for various practical purposes. In this study, we tried discerning data analysis for clarify the chemical action to the ionotoropic glutamate receptor (GluR), whereby an ion-flux assay of post-synapse Model Cells is performed and are analyzed based on a chemometrics approach. The dynamic behavior of the GluR of post-synapse Model Cell was assayed with multivariate data analysis methods namely hierarchical cluster analysis (HCA) and principal component analysis (PCA). By using HCA, we can identify and remove the non-responding samples. By using PCA, the effect of chemicals on the dynamic behavior of ion flux through GluR can be recognized clearly; as either agonist or antagonist. As shown in the results, the GluR-based assay by post-synapse Model Cell with data analysis methods provide a sodium influx profile which is derived by an agonists or antagonists application. By employing the data analysis methods, PCA and HCA, it is possible to develop a smart Cellular biosensing system for qualified analysis.
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engineered synapse Model Cell genetic construction and chemical evaluation for reproducible high throughput analysis
Analytical and Bioanalytical Chemistry, 2010Co-Authors: Satoshi Migita, Akito Tateishi, Kari Keinanen, Tetsuya HaruyamaAbstract:Bioassay Models of neural functions must lend themselves to high-throughput analysis in neural drug discovery. However, smart analysis methods for these functions have not yet been fully established. Here, we describe the development of a synapse Model for Cell-based biosensing. The engineered synapse Model Cell expresses ionotropic glutamate receptor on its surface, like the neural postsynaptic membrane. The advantages of the Model Cell are the ease of handling and reproducibility as compared with the cultured neural Cell, and it can be employed to evaluate receptor function through ion flux analysis. The agonist-induced sodium influx was monitored as an agonist concentration-dependent increase in the observed fluorescence signal. Furthermore, we found that our Model Cell enables the correction of uneven Cellular signal levels using a reporter system. Our engineered synapse Model Cell can be employed as a powerful tool for the screening of lead substances in pharmaceutical high-throughput analysis.
Feng Wei - One of the best experts on this subject based on the ideXlab platform.
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Observing a Model Ion Channel Gating Action in Model Cell Membranes in Real Time in Situ: Membrane Potential Change Induced Alamethicin Orientation Change
2016Co-Authors: Feng Wei, Pei Yang, Joshua Jasensky, Andrew P. Boughton, Zhan ChenAbstract:Ion channels play crucial roles in transport and regulatory functions of living Cells. Understanding the gating mechanisms of these channels is important to understanding and treating diseases that have been linked to ion channels. One potential Model peptide for studying the mechanism of ion channel gating is alamethicin, which adopts a split α/310-helix structure and responds to changes in electric potential. In this study, sum frequency generation vibrational spectroscopy (SFG-VS), supplemented by attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR), has been applied to characterize interactions between alamethicin (a Model for larger channel proteins) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) lipid bilayers in the presence of an electric potential across the membrane. The membrane potential difference was controlled by changing the pH of the solution in contact with the bilayer and was measured using fluorescence spectroscopy. The orientation angle of alamethicin in POPC lipid bilayers was then determined at different pH values using polarized SFG amide I spectra. Assuming that all molecules adopt the same orientation (a δ distribution), at pH = 6.7 the α-helix at the N-terminus and the 310-helix at the C-terminus tilt at about 72° (θ1) and 50° (θ2) versus the surface normal, respectively. When pH increases to 11.9, θ1 and θ2 decrease to 56.5° and 45°, respectively. The δ distribution assumption was verified using a combination of SFG and ATR-FTIR measurements, which showed a quite narrow distribution in the angle of θ1 for both pH conditions. This indicates that all alamethicin molecules at the surface adopt a nearly identical orientation in POPC lipid bilayers. The localized pH change in proximity to the bilayer modulates the membrane potential and thus induces a decrease in both the tilt and the bend angles of the two helices in alamethicin. This is the first reported application of SFG to the study of Model ion channel gating mechanisms in Model Cell membranes
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observing a Model ion channel gating action in Model Cell membranes in real time in situ membrane potential change induced alamethicin orientation change
Journal of the American Chemical Society, 2012Co-Authors: Feng Wei, Pei Yang, Joshua Jasensky, Andrew P Boughton, Zhan ChenAbstract:Ion channels play crucial roles in transport and regulatory functions of living Cells. Understanding the gating mechanisms of these channels is important to understanding and treating diseases that have been linked to ion channels. One potential Model peptide for studying the mechanism of ion channel gating is alamethicin, which adopts a split α/3(10)-helix structure and responds to changes in electric potential. In this study, sum frequency generation vibrational spectroscopy (SFG-VS), supplemented by attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR), has been applied to characterize interactions between alamethicin (a Model for larger channel proteins) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) lipid bilayers in the presence of an electric potential across the membrane. The membrane potential difference was controlled by changing the pH of the solution in contact with the bilayer and was measured using fluorescence spectroscopy. The orientation angle of alamethicin in POPC lipid bilayers was then determined at different pH values using polarized SFG amide I spectra. Assuming that all molecules adopt the same orientation (a δ distribution), at pH = 6.7 the α-helix at the N-terminus and the 3(10)-helix at the C-terminus tilt at about 72° (θ(1)) and 50° (θ(2)) versus the surface normal, respectively. When pH increases to 11.9, θ(1) and θ(2) decrease to 56.5° and 45°, respectively. The δ distribution assumption was verified using a combination of SFG and ATR-FTIR measurements, which showed a quite narrow distribution in the angle of θ(1) for both pH conditions. This indicates that all alamethicin molecules at the surface adopt a nearly identical orientation in POPC lipid bilayers. The localized pH change in proximity to the bilayer modulates the membrane potential and thus induces a decrease in both the tilt and the bend angles of the two helices in alamethicin. This is the first reported application of SFG to the study of Model ion channel gating mechanisms in Model Cell membranes.
