Amy Won - Academia.edu (original) (raw)

Papers by Amy Won

Research paper thumbnail of Postalkylation of a Common mPEG-b-PAGE Precursor to Produce Tunable Morphologies of Spheres, Filomicelles, Disks, and Polymersomes

Research paper thumbnail of Mapping Local Nanoscale Changes in Cell Tension and Stiffness by Combinatorial Microscopies

Biophysical Journal, 2014

Research paper thumbnail of Binding of TDP-43 to the 3'UTR of its cognate mRNA enhances its solubility

Biochemistry, Jan 23, 2014

TAR DNA binding protein of 43 kDa (TDP-43) has been implicated in the pathogenesis of a broad ran... more TAR DNA binding protein of 43 kDa (TDP-43) has been implicated in the pathogenesis of a broad range of neurodegenerative diseases termed TDP-43 proteinopathies, which encompass a spectrum of diseases ranging from amyotrophic lateral sclerosis to frontotemporal dementia. Pathologically misfolded and aggregated forms of TDP-43 are found in cytoplasmic inclusion bodies of affected neurons in these diseases. The mechanism by which TDP-43 misfolding causes disease is not well-understood. Current hypotheses postulate that the TDP-43 aggregation process plays a major role in pathogenesis. We amplify that hypothesis and suggest that binding of cognate ligands to TDP-43 can stabilize the native functional state of the protein and ameliorate aggregation. We expressed recombinant TDP-43 containing an N-terminal Venus yellow fluorescent protein tag in Escherichia coli and induced its aggregation by altering solvent salt concentrations and examined the extent to which various oligonucleotide mol...

Research paper thumbnail of Maps and Floorplans

Research paper thumbnail of Continuing education courses

Research paper thumbnail of Tracking Ceacam1 Interactions and Dynamics with homo-FRET and Image Correlation Techniques

Biophysical Journal, 2015

Research paper thumbnail of Interactome Analyses Identify Ties of PrPC and Its Mammalian Paralogs to Oligomannosidic N-Glycans and Endoplasmic Reticulum-Derived Chaperones

PLoS Pathogens, 2009

The physiological environment which hosts the conformational conversion of the cellular prion pro... more The physiological environment which hosts the conformational conversion of the cellular prion protein (PrP C ) to diseaseassociated isoforms has remained enigmatic. A quantitative investigation of the PrP C interactome was conducted in a cell culture model permissive to prion replication. To facilitate recognition of relevant interactors, the study was extended to Doppel (Prnd) and Shadoo (Sprn), two mammalian PrP C paralogs. Interestingly, this work not only established a similar physiological environment for the three prion protein family members in neuroblastoma cells, but also suggested direct interactions amongst them. Furthermore, multiple interactions between PrP C and the neural cell adhesion molecule, the laminin receptor precursor, Na/K ATPases and protein disulfide isomerases (PDI) were confirmed, thereby reconciling previously separate findings. Subsequent validation experiments established that interactions of PrP C with PDIs may extend beyond the endoplasmic reticulum and may play a hitherto unrecognized role in the accumulation of PrP Sc . A simple hypothesis is presented which accounts for the majority of interactions observed in uninfected cells and suggests that PrP C organizes its molecular environment on account of its ability to bind to adhesion molecules harboring immunoglobulin-like domains, which in turn recognize oligomannose-bearing membrane proteins.

Research paper thumbnail of The effects of L- to D-isomerization and C-terminus deamidation on the secondary structure of antimicrobial peptide Anoplin in aqueous and membrane mimicking environment

Journal of Raman Spectroscopy, 2010

UV resonance Raman spectra of the antimicrobial peptide (AMP) Anoplin (L-Anoplin-NH 2 ) and two o... more UV resonance Raman spectra of the antimicrobial peptide (AMP) Anoplin (L-Anoplin-NH 2 ) and two of its derivatives (enantiomer D-Anoplin-NH 2 and C-terminus deamidated L-Anoplin-OH) were measured in aqueous buffer solution and in membrane-mimetic environments including 2,2,2-trifluoro ethanol (TFE), zwitterionic lipid dipalmitoylglycerophosphocholine (DPPC) and anionic lipid dipalmitoylglycerophosphoglycerol (DPPG) vesicle solutions. All three peptides were found to adopt random-coil/β turn-like conformation in aqueous solution over the temperature range of 1-60 • C. The conformation was found to become more α-helical in membrane-mimetic solutions such as TFE and DPPG but not in DPPC for all Anoplin derivatives. The data demonstrate that Anoplin preferentially binds to the anionic over the zwitterionic model cell membranes. Results also showed that deamidation does not change the conformation of L-Ano-NH 2 very significantly, but does alter membrane rupturing and antimicrobial activities thus confirming that it is the physicochemical properties rather than the peptide conformation that define the mechanism of AMP action.

Research paper thumbnail of Imaging interactions of cationic antimicrobial peptides with model lipid monolayers using X-ray spectromicroscopy

European Biophysics Journal, 2011

The interaction of antimicrobial peptide anoplin with 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(... more The interaction of antimicrobial peptide anoplin with 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] lipid monolayers was imaged with atomic force microscopy, scanning transmission X-ray microscopy, and X-ray photoemission electron microscopy. X-ray absorption spectromicroscopy of the surface revealed the domains of the phase-segregated surface to be composed of 98(±5)% lipid while the matrix consisted of a ~50:50 lipid-peptide mixture. We show X-ray spectromicroscopy to be a valuable quantitative tool for label-free imaging of lipid monolayers with antimicrobial peptides at a lateral spatial resolution below 80 nm.

Research paper thumbnail of Investigating the effect of a single glycine to alanine substitution on interactions of antimicrobial peptide latarcin 2a with a lipid membrane

European Biophysics Journal, 2011

Latarcins are linear, a-helical antimicrobial peptides purified from the venom of the Central Asi... more Latarcins are linear, a-helical antimicrobial peptides purified from the venom of the Central Asian spider Lachesana tarabaevi, with lytic activity against Gram-positive and Gram-negative bacteria, erythrocytes, and yeast at micromolar concentrations. In this work, we investigated the role of the hinge in latarcin 2a (ltc2a, GLFGKLIKKFGRKAISYAVKKARGKH-COOH), which adopts a helix-hinge-helix conformation in membranemimicking environments, on peptide-membrane interactions and its potential effect on the selective toxicity of the peptide. A modified latarcin 2a, ltc2aG11A, obtained by replacing the glycine at position 11 with alanine (ltc2aG11A, GLFGKL IKKFARKAISYAVKKARGKH-COOH), adopts a more rigid structure due to the reduced conformational flexibility. Langmuir monolayer measurements combined with atomic force microscopy and X-ray photoemission electron microscopy (X-PEEM) indicate that both peptides bind and insert preferentially into anionic compared with zwitterionic phospholipid monolayers. Modified ltc2aG11A was found to be more disruptive of supported phospholipid bilayer modeling mammalian cell membrane. However, no considerable difference in lytic activity of the two peptides toward bacterial membrane was found. Overall the data indicate that decrease in the flexibility of ltc2a induced by the modification in the hinge region is likely to increase the peptide's nonspecific interactions with zwitterionic cell membranes and potentially increase its toxicity against eukaryotic cells.

Research paper thumbnail of Interaction Between Lytic Peptide Latarcin 2A and Supported Lipid Bilayers Studied by in Situ Atomic Force Microscopy

Biophysical Journal, 2012

Research paper thumbnail of Interactions of Antimicrobial Peptide Latarcin With Model Cell Membrane

Biophysical Journal, 2010

Research paper thumbnail of The Effect of Point Mutations on the Secondary Structure and Membrane Interaction of Antimicrobial Peptide Anoplin

Biophysical Journal, 2011

Research paper thumbnail of Deamidation Weakens Membrane Binding Properties of Antimicrobial Peptide Anoplin

Biophysical Journal, 2009

Research paper thumbnail of Molecular Mechanism of pEM-2 Activity

Biophysical Journal, 2009

Research paper thumbnail of Effect of L- to D-Peptide Isomerisation on the Activity of Antimicrobial Peptide Anoplin

Biophysical Journal, 2010

water-soluble polymer, PEG, to the lamellar phase of DMPC or of several surfactants, induces a to... more water-soluble polymer, PEG, to the lamellar phase of DMPC or of several surfactants, induces a topological transformation to a vesicular phase. Such transition can be understood in terms of a modification of the elastic properties of the membranes. In this work we perform a dynamic light scattering (DLS) study of the effect of PEG on phospholipid and surfactant bilayers. The experimental results show that the addition of the polymer slows down the dynamics of the membranes. From fits to an available theory for the scattering of dilute membrane systems, we have calculated the bending elastic modulus of the bilayers. This modulus increases with increasing polymer concentration, thus confirming that the macromolecule modifies the elastic properties of the membranes.

Research paper thumbnail of Imaging the membrane lytic activity of bioactive peptide latarcin 2a

Biochimica et Biophysica Acta (BBA) - Biomembranes, 2012

Latarcin 2a (ltc2a, GLFGKLIKKFGRKAISYAVKKARGKH-COOH) is a short linear antimicrobial and cytolyti... more Latarcin 2a (ltc2a, GLFGKLIKKFGRKAISYAVKKARGKH-COOH) is a short linear antimicrobial and cytolytic peptide extracted from the venom of the Central Asian spider, Lachesana tarabaevi, with lytic activity against Gram-positive and Gram-negative bacteria, erythrocytes, and yeast at micromolar concentrations. Ltc2a adopts a helix-hinge-helix structure in membrane mimicking environment, whereas its derivative latarcin 2aG11A (ltc2aG11A, GLFGKLIKKFARKAISYAVKKARGKH-COOH), likely adopts a more rigid structure, demonstrates stronger nonspecific interaction with the zwitterionic membrane, and is potentially more toxic against eukaryotic cells. In this work, interactions of these two ltc2a derivatives with supported "raft" lipid bilayer (1,2-dioleoyl-sn-glycero-3-phosphocholin/egg sphingomyelin/cholesterol 40/40/20mol%) were studied by in situ atomic force microscopy in order to investigate the potential anticancer activity of the peptides since some breast and prostate cancer cell lines contain higher levels of cholesterol-rich lipid rafts than non-cancer cells. Both peptides induced reorganization of the raft model membrane by reducing line tension of the liquid ordered phase. Ltc2aG11A induced membrane thinning likely due to membrane interdigitation. Formation of large pores by the peptides in the bilayer was observed. Cholesterol was found to attenuate membrane disruption by the peptides. Finally, leakage assay showed that both peptides have similar membrane permeability toward various model membrane vesicles.

Research paper thumbnail of Interactions of antimicrobial peptide from C-terminus of myotoxin II with phospholipid mono- and bilayers

Biochimica et Biophysica Acta (BBA) - Biomembranes, 2009

Comparative studies of the effect of a short synthetic cationic peptide, pEM-2 (KKWRWWLKALAKK), d... more Comparative studies of the effect of a short synthetic cationic peptide, pEM-2 (KKWRWWLKALAKK), derived from the C-terminus of myotoxin II from the venom of the snake Bothrops asper on phospholipid mono-and bilayers were performed by means of Langmuir Blodgett (LB) monolayer technique, atomic force microscopy and calcein leakage assay. Phospholipid mono-and bilayers composed of single zwitterionic or anionic phospholipids as well as lipid mixtures mimicking bacterial cell membrane were used. LB measurements indicate that the peptide binds to both anionic and zwitterionic phospholipid monolayers at low surface pressure but only to anionic at high surface pressure. Preferential interaction of the peptide with anionic phospholipid monolayer is also supported by a more pronounced change of the monolayer pressure/area isotherms induced by the peptide. AFM imaging reveals the presence of nanoscale aggregates in lipid/ peptide mixture monolayers. At the same time, calcein leakage experiment demonstrated that pEM-2 induces stronger disruption of zwitterionic than anionic bilayers. Results of the study indicate that electrostatic interactions play a significant role in the initial recognition and binding of pEM-2 to the cell membrane. However, membrane rupturing activity of the peptide depends on interactions other than simple ionic attraction.

Research paper thumbnail of Investigating the effects of L- to D-amino acid substitution and deamidation on the activity and membrane interactions of antimicrobial peptide anoplin

Biochimica et Biophysica Acta (BBA) - Biomembranes, 2011

Isolated from the venom sac of solitary spider wasp, Anoplius samariensis, anoplin is the smalles... more Isolated from the venom sac of solitary spider wasp, Anoplius samariensis, anoplin is the smallest linear α-helical antimicrobial peptide found naturally with broad spectrum activity against both Gram-positive and Gram-negative bacteria, and little hemolytic activity toward human erythrocytes. Deamidation was found to decrease the peptide's antibacterial properties. In the present work, interactions of amidated (Ano-NH2) and deamidated (Ano-OH) forms of anoplin as well as Ano-NH2 composed of all D-amino acids (D-Ano-NH2) with model cell membranes were investigated by means of Langmuir Blodgett (LB) technique, atomic force microscopy (AFM), X-ray photoemission electron microscopy (X-PEEM) and carboxyfluorescein leakage assay in order to gain a better understanding of the effect of these peptide modifications on membrane binding and lytic properties. According to LB, all three peptides form stable monolayers at the air/water interface with Ano-NH2 occupying a slightly greater area per molecule than Ano-OH. All three forms of the peptide interact preferentially with anionic 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (DPPG), rather than zwitterionic 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) lipid monolayer. Peptides form nanoscale clusters in zwitterionic but not in anionic monolayers. Finally, membrane lytic activity of all derivatives was found to depend strongly on membrane composition and lipid/peptide ratio. The results suggest that amidated forms of peptides are likely to possess higher membrane binding affinity due to the increased charge.

Research paper thumbnail of Mitochondrial Genome Maintenance 1 (Mgm1) Protein Alters Membrane Topology and Promotes Local Membrane Bending

Journal of Molecular Biology, 2015

Large GTPases of the dynamin superfamily promote membrane fusion and division, processes that are... more Large GTPases of the dynamin superfamily promote membrane fusion and division, processes that are crucial for intracellular trafficking and organellar dynamics. To promote membrane scission, dynamin proteins polymerize, wrap around, and constrict the membrane; however, the mechanism underlying their role in membrane fusion remains unclear. We previously reported that the mitochondrial dynamin-related protein mitochondrial genome maintenance 1 (Mgm1) mediates fusion by first tethering opposing membranes and then undergoing a nucleotide-dependent structural transition. However, it is still unclear how Mgm1 directly affects the membrane to drive fusion of tethered membranes. Here, we show that Mgm1 association with the membrane alters the topography of the membrane, promoting local membrane bending. We also demonstrate that Mgm1 creates membrane ruffles resulting in the formation of tubular structures on both supported lipid bilayers and liposomes. These data suggest that Mgm1 membrane interactions impose a mechanical force on the membrane to overcome the hydrophilic repulsion of the phospholipid head groups and initiate the fusion reaction. The work reported here provides new insights into a possible mechanism of Mgm1-driven mitochondrial membrane fusion and sheds light into how members of the dynamin superfamily function as fusion molecules.

Research paper thumbnail of Postalkylation of a Common mPEG-b-PAGE Precursor to Produce Tunable Morphologies of Spheres, Filomicelles, Disks, and Polymersomes

Research paper thumbnail of Mapping Local Nanoscale Changes in Cell Tension and Stiffness by Combinatorial Microscopies

Biophysical Journal, 2014

Research paper thumbnail of Binding of TDP-43 to the 3'UTR of its cognate mRNA enhances its solubility

Biochemistry, Jan 23, 2014

TAR DNA binding protein of 43 kDa (TDP-43) has been implicated in the pathogenesis of a broad ran... more TAR DNA binding protein of 43 kDa (TDP-43) has been implicated in the pathogenesis of a broad range of neurodegenerative diseases termed TDP-43 proteinopathies, which encompass a spectrum of diseases ranging from amyotrophic lateral sclerosis to frontotemporal dementia. Pathologically misfolded and aggregated forms of TDP-43 are found in cytoplasmic inclusion bodies of affected neurons in these diseases. The mechanism by which TDP-43 misfolding causes disease is not well-understood. Current hypotheses postulate that the TDP-43 aggregation process plays a major role in pathogenesis. We amplify that hypothesis and suggest that binding of cognate ligands to TDP-43 can stabilize the native functional state of the protein and ameliorate aggregation. We expressed recombinant TDP-43 containing an N-terminal Venus yellow fluorescent protein tag in Escherichia coli and induced its aggregation by altering solvent salt concentrations and examined the extent to which various oligonucleotide mol...

Research paper thumbnail of Maps and Floorplans

Research paper thumbnail of Continuing education courses

Research paper thumbnail of Tracking Ceacam1 Interactions and Dynamics with homo-FRET and Image Correlation Techniques

Biophysical Journal, 2015

Research paper thumbnail of Interactome Analyses Identify Ties of PrPC and Its Mammalian Paralogs to Oligomannosidic N-Glycans and Endoplasmic Reticulum-Derived Chaperones

PLoS Pathogens, 2009

The physiological environment which hosts the conformational conversion of the cellular prion pro... more The physiological environment which hosts the conformational conversion of the cellular prion protein (PrP C ) to diseaseassociated isoforms has remained enigmatic. A quantitative investigation of the PrP C interactome was conducted in a cell culture model permissive to prion replication. To facilitate recognition of relevant interactors, the study was extended to Doppel (Prnd) and Shadoo (Sprn), two mammalian PrP C paralogs. Interestingly, this work not only established a similar physiological environment for the three prion protein family members in neuroblastoma cells, but also suggested direct interactions amongst them. Furthermore, multiple interactions between PrP C and the neural cell adhesion molecule, the laminin receptor precursor, Na/K ATPases and protein disulfide isomerases (PDI) were confirmed, thereby reconciling previously separate findings. Subsequent validation experiments established that interactions of PrP C with PDIs may extend beyond the endoplasmic reticulum and may play a hitherto unrecognized role in the accumulation of PrP Sc . A simple hypothesis is presented which accounts for the majority of interactions observed in uninfected cells and suggests that PrP C organizes its molecular environment on account of its ability to bind to adhesion molecules harboring immunoglobulin-like domains, which in turn recognize oligomannose-bearing membrane proteins.

Research paper thumbnail of The effects of L- to D-isomerization and C-terminus deamidation on the secondary structure of antimicrobial peptide Anoplin in aqueous and membrane mimicking environment

Journal of Raman Spectroscopy, 2010

UV resonance Raman spectra of the antimicrobial peptide (AMP) Anoplin (L-Anoplin-NH 2 ) and two o... more UV resonance Raman spectra of the antimicrobial peptide (AMP) Anoplin (L-Anoplin-NH 2 ) and two of its derivatives (enantiomer D-Anoplin-NH 2 and C-terminus deamidated L-Anoplin-OH) were measured in aqueous buffer solution and in membrane-mimetic environments including 2,2,2-trifluoro ethanol (TFE), zwitterionic lipid dipalmitoylglycerophosphocholine (DPPC) and anionic lipid dipalmitoylglycerophosphoglycerol (DPPG) vesicle solutions. All three peptides were found to adopt random-coil/β turn-like conformation in aqueous solution over the temperature range of 1-60 • C. The conformation was found to become more α-helical in membrane-mimetic solutions such as TFE and DPPG but not in DPPC for all Anoplin derivatives. The data demonstrate that Anoplin preferentially binds to the anionic over the zwitterionic model cell membranes. Results also showed that deamidation does not change the conformation of L-Ano-NH 2 very significantly, but does alter membrane rupturing and antimicrobial activities thus confirming that it is the physicochemical properties rather than the peptide conformation that define the mechanism of AMP action.

Research paper thumbnail of Imaging interactions of cationic antimicrobial peptides with model lipid monolayers using X-ray spectromicroscopy

European Biophysics Journal, 2011

The interaction of antimicrobial peptide anoplin with 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(... more The interaction of antimicrobial peptide anoplin with 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] lipid monolayers was imaged with atomic force microscopy, scanning transmission X-ray microscopy, and X-ray photoemission electron microscopy. X-ray absorption spectromicroscopy of the surface revealed the domains of the phase-segregated surface to be composed of 98(±5)% lipid while the matrix consisted of a ~50:50 lipid-peptide mixture. We show X-ray spectromicroscopy to be a valuable quantitative tool for label-free imaging of lipid monolayers with antimicrobial peptides at a lateral spatial resolution below 80 nm.

Research paper thumbnail of Investigating the effect of a single glycine to alanine substitution on interactions of antimicrobial peptide latarcin 2a with a lipid membrane

European Biophysics Journal, 2011

Latarcins are linear, a-helical antimicrobial peptides purified from the venom of the Central Asi... more Latarcins are linear, a-helical antimicrobial peptides purified from the venom of the Central Asian spider Lachesana tarabaevi, with lytic activity against Gram-positive and Gram-negative bacteria, erythrocytes, and yeast at micromolar concentrations. In this work, we investigated the role of the hinge in latarcin 2a (ltc2a, GLFGKLIKKFGRKAISYAVKKARGKH-COOH), which adopts a helix-hinge-helix conformation in membranemimicking environments, on peptide-membrane interactions and its potential effect on the selective toxicity of the peptide. A modified latarcin 2a, ltc2aG11A, obtained by replacing the glycine at position 11 with alanine (ltc2aG11A, GLFGKL IKKFARKAISYAVKKARGKH-COOH), adopts a more rigid structure due to the reduced conformational flexibility. Langmuir monolayer measurements combined with atomic force microscopy and X-ray photoemission electron microscopy (X-PEEM) indicate that both peptides bind and insert preferentially into anionic compared with zwitterionic phospholipid monolayers. Modified ltc2aG11A was found to be more disruptive of supported phospholipid bilayer modeling mammalian cell membrane. However, no considerable difference in lytic activity of the two peptides toward bacterial membrane was found. Overall the data indicate that decrease in the flexibility of ltc2a induced by the modification in the hinge region is likely to increase the peptide's nonspecific interactions with zwitterionic cell membranes and potentially increase its toxicity against eukaryotic cells.

Research paper thumbnail of Interaction Between Lytic Peptide Latarcin 2A and Supported Lipid Bilayers Studied by in Situ Atomic Force Microscopy

Biophysical Journal, 2012

Research paper thumbnail of Interactions of Antimicrobial Peptide Latarcin With Model Cell Membrane

Biophysical Journal, 2010

Research paper thumbnail of The Effect of Point Mutations on the Secondary Structure and Membrane Interaction of Antimicrobial Peptide Anoplin

Biophysical Journal, 2011

Research paper thumbnail of Deamidation Weakens Membrane Binding Properties of Antimicrobial Peptide Anoplin

Biophysical Journal, 2009

Research paper thumbnail of Molecular Mechanism of pEM-2 Activity

Biophysical Journal, 2009

Research paper thumbnail of Effect of L- to D-Peptide Isomerisation on the Activity of Antimicrobial Peptide Anoplin

Biophysical Journal, 2010

water-soluble polymer, PEG, to the lamellar phase of DMPC or of several surfactants, induces a to... more water-soluble polymer, PEG, to the lamellar phase of DMPC or of several surfactants, induces a topological transformation to a vesicular phase. Such transition can be understood in terms of a modification of the elastic properties of the membranes. In this work we perform a dynamic light scattering (DLS) study of the effect of PEG on phospholipid and surfactant bilayers. The experimental results show that the addition of the polymer slows down the dynamics of the membranes. From fits to an available theory for the scattering of dilute membrane systems, we have calculated the bending elastic modulus of the bilayers. This modulus increases with increasing polymer concentration, thus confirming that the macromolecule modifies the elastic properties of the membranes.

Research paper thumbnail of Imaging the membrane lytic activity of bioactive peptide latarcin 2a

Biochimica et Biophysica Acta (BBA) - Biomembranes, 2012

Latarcin 2a (ltc2a, GLFGKLIKKFGRKAISYAVKKARGKH-COOH) is a short linear antimicrobial and cytolyti... more Latarcin 2a (ltc2a, GLFGKLIKKFGRKAISYAVKKARGKH-COOH) is a short linear antimicrobial and cytolytic peptide extracted from the venom of the Central Asian spider, Lachesana tarabaevi, with lytic activity against Gram-positive and Gram-negative bacteria, erythrocytes, and yeast at micromolar concentrations. Ltc2a adopts a helix-hinge-helix structure in membrane mimicking environment, whereas its derivative latarcin 2aG11A (ltc2aG11A, GLFGKLIKKFARKAISYAVKKARGKH-COOH), likely adopts a more rigid structure, demonstrates stronger nonspecific interaction with the zwitterionic membrane, and is potentially more toxic against eukaryotic cells. In this work, interactions of these two ltc2a derivatives with supported "raft" lipid bilayer (1,2-dioleoyl-sn-glycero-3-phosphocholin/egg sphingomyelin/cholesterol 40/40/20mol%) were studied by in situ atomic force microscopy in order to investigate the potential anticancer activity of the peptides since some breast and prostate cancer cell lines contain higher levels of cholesterol-rich lipid rafts than non-cancer cells. Both peptides induced reorganization of the raft model membrane by reducing line tension of the liquid ordered phase. Ltc2aG11A induced membrane thinning likely due to membrane interdigitation. Formation of large pores by the peptides in the bilayer was observed. Cholesterol was found to attenuate membrane disruption by the peptides. Finally, leakage assay showed that both peptides have similar membrane permeability toward various model membrane vesicles.

Research paper thumbnail of Interactions of antimicrobial peptide from C-terminus of myotoxin II with phospholipid mono- and bilayers

Biochimica et Biophysica Acta (BBA) - Biomembranes, 2009

Comparative studies of the effect of a short synthetic cationic peptide, pEM-2 (KKWRWWLKALAKK), d... more Comparative studies of the effect of a short synthetic cationic peptide, pEM-2 (KKWRWWLKALAKK), derived from the C-terminus of myotoxin II from the venom of the snake Bothrops asper on phospholipid mono-and bilayers were performed by means of Langmuir Blodgett (LB) monolayer technique, atomic force microscopy and calcein leakage assay. Phospholipid mono-and bilayers composed of single zwitterionic or anionic phospholipids as well as lipid mixtures mimicking bacterial cell membrane were used. LB measurements indicate that the peptide binds to both anionic and zwitterionic phospholipid monolayers at low surface pressure but only to anionic at high surface pressure. Preferential interaction of the peptide with anionic phospholipid monolayer is also supported by a more pronounced change of the monolayer pressure/area isotherms induced by the peptide. AFM imaging reveals the presence of nanoscale aggregates in lipid/ peptide mixture monolayers. At the same time, calcein leakage experiment demonstrated that pEM-2 induces stronger disruption of zwitterionic than anionic bilayers. Results of the study indicate that electrostatic interactions play a significant role in the initial recognition and binding of pEM-2 to the cell membrane. However, membrane rupturing activity of the peptide depends on interactions other than simple ionic attraction.

Research paper thumbnail of Investigating the effects of L- to D-amino acid substitution and deamidation on the activity and membrane interactions of antimicrobial peptide anoplin

Biochimica et Biophysica Acta (BBA) - Biomembranes, 2011

Isolated from the venom sac of solitary spider wasp, Anoplius samariensis, anoplin is the smalles... more Isolated from the venom sac of solitary spider wasp, Anoplius samariensis, anoplin is the smallest linear α-helical antimicrobial peptide found naturally with broad spectrum activity against both Gram-positive and Gram-negative bacteria, and little hemolytic activity toward human erythrocytes. Deamidation was found to decrease the peptide's antibacterial properties. In the present work, interactions of amidated (Ano-NH2) and deamidated (Ano-OH) forms of anoplin as well as Ano-NH2 composed of all D-amino acids (D-Ano-NH2) with model cell membranes were investigated by means of Langmuir Blodgett (LB) technique, atomic force microscopy (AFM), X-ray photoemission electron microscopy (X-PEEM) and carboxyfluorescein leakage assay in order to gain a better understanding of the effect of these peptide modifications on membrane binding and lytic properties. According to LB, all three peptides form stable monolayers at the air/water interface with Ano-NH2 occupying a slightly greater area per molecule than Ano-OH. All three forms of the peptide interact preferentially with anionic 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (DPPG), rather than zwitterionic 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) lipid monolayer. Peptides form nanoscale clusters in zwitterionic but not in anionic monolayers. Finally, membrane lytic activity of all derivatives was found to depend strongly on membrane composition and lipid/peptide ratio. The results suggest that amidated forms of peptides are likely to possess higher membrane binding affinity due to the increased charge.

Research paper thumbnail of Mitochondrial Genome Maintenance 1 (Mgm1) Protein Alters Membrane Topology and Promotes Local Membrane Bending

Journal of Molecular Biology, 2015

Large GTPases of the dynamin superfamily promote membrane fusion and division, processes that are... more Large GTPases of the dynamin superfamily promote membrane fusion and division, processes that are crucial for intracellular trafficking and organellar dynamics. To promote membrane scission, dynamin proteins polymerize, wrap around, and constrict the membrane; however, the mechanism underlying their role in membrane fusion remains unclear. We previously reported that the mitochondrial dynamin-related protein mitochondrial genome maintenance 1 (Mgm1) mediates fusion by first tethering opposing membranes and then undergoing a nucleotide-dependent structural transition. However, it is still unclear how Mgm1 directly affects the membrane to drive fusion of tethered membranes. Here, we show that Mgm1 association with the membrane alters the topography of the membrane, promoting local membrane bending. We also demonstrate that Mgm1 creates membrane ruffles resulting in the formation of tubular structures on both supported lipid bilayers and liposomes. These data suggest that Mgm1 membrane interactions impose a mechanical force on the membrane to overcome the hydrophilic repulsion of the phospholipid head groups and initiate the fusion reaction. The work reported here provides new insights into a possible mechanism of Mgm1-driven mitochondrial membrane fusion and sheds light into how members of the dynamin superfamily function as fusion molecules.