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| 2. |
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| 3. |
- Ali Doosti, Baharan, 1991, et al.
(författare)
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Ca2+ Gradient Induces Membrane Bending and Formation of Nanotubes in Giant Lipid Vesicles
- 2016
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Ingår i: Biophysical Journal. - : Elsevier BV. - 0006-3495 .- 1542-0086. ; 110:3, s. 584A-584A
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Tidskriftsartikel (refereegranskat)abstract
- Reshaping and bending of the cell membrane is imperative in many processes such as cell division, filopodia formation, and endocytosis. Understanding these shape transitions, will help to elucidate the underlying mechanisms of these essential cellular processes. In our work, we investigate an interplay between cell membrane morphology and chemical stimulation by constructing a biomimetic model system. More specifically, giant lipid vesicles were exposed to a chemical gradient of Ca2+, which was established over the membrane surface.
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| 4. |
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| 5. |
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| 6. |
- Alizadehheidari, Mohammadreza, 1987, et al.
(författare)
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Unfolding of nanoconfined circular DNA
- 2015
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Ingår i: BIOPHYSICAL JOURNAL. - : Elsevier BV. - 0006-3495 .- 1542-0086. ; 108:2 Supplement 1
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)
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| 7. |
- Almaqwashi, A. A., et al.
(författare)
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Dissecting the Dynamic Pathways of Stereoselective DNA Threading Intercalation
- 2016
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Ingår i: Biophysical Journal. - : Elsevier BV. - 0006-3495 .- 1542-0086. ; 110:6, s. 1255-1263
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Tidskriftsartikel (refereegranskat)abstract
- DNA intercalators that have high affinity and slow kinetics are developed for potential DNA-targeted therapeutics. Although many natural intercalators contain multiple chiral subunits, only intercalators with a single chiral unit have been quantitatively probed. Dumbbell-shaped DNA threading intercalators represent the next order of structural complexity relative to simple intercalators, and can provide significant insights into the stereoselectivity of DNA-ligand intercalation. We investigated DNA threading intercalation by binuclear ruthenium complex [mu-dppzip(phen)(4)Ru-2](4+) (Piz). Four Piz stereoisomers are defined by the chirality of the intercalating subunit (Ru(phen)(2)dppz) and the distal subunit (Ru(phen)(2)ip), respectively, each of which can be either right-handed (Delta) or left-handed (Lambda). We used optical tweezers to measure single DNA molecule elongation due to threading intercalation, revealing force-dependent DNA intercalation rates and equilibrium dissociation constants. The force spectroscopy analysis provided the zero-force DNA binding affinity, the equilibrium DNA-ligand elongation Delta x(eq), and the dynamic DNA structural deformations during ligand association x(on) and dissociation x(off). We found that Piz stereoisomers exhibit over 20-fold differences in DNA binding affinity, from a K-d of 27 +/- 3 nM for (Delta,Lambda)-Piz to a K-d of 622 +/- 55 nM for (Lambda,Delta)-Piz. The striking affinity decrease is correlated with increasing Delta x(eq) from 0.30 +/- 0.02 to 0.48 +/- 0.02 nm and x(on) from 0.25 +/- 0.01 to 0.46 +/- 0.02 nm, but limited x(off) changes. Notably, the affinity and threading kinetics is 10-fold enhanced for right-handed intercalating subunits, and 2- to 5-fold enhanced for left-handed distal subunits. These findings demonstrate sterically dispersed transition pathways and robust DNA structural recognition of chiral intercalators, which are critical for optimizing DNA binding affinity and kinetics.
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| 13. |
- Badell, Maria Valldeperas, et al.
(författare)
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Lipid Sponge-Phase Nanoparticles as Carriers for Enzymes
- 2018
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Ingår i: Biophysical Journal. - : Cell Press. - 0006-3495 .- 1542-0086. ; 114:3, suppl 1, s. 15A-15A
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- Immobilization of enzymes into different support materials has been widely studied as means to control their activity and stability. Here we will consider lipid liquid crystalline phases as enzyme carriers, as they have been demonstrated to have a high potential in a range of applications such as drug delivery, protein encapsulation or crystallization thanks to the wide range of self-assembly structures they can form, which have cavities of nano-scale dimensions. Furthermore, such structures have also been observed in a range of living organisms. Although, reverse cubic or hexagonal lipid aqueous phase can be used to entrap smaller biomolecules, it is still challenging to encapsulate bioactive macromolecules, such as proteins. Here, we will present a novel lipid system able to form highly swollen sponge phases (L3), with aqueous pores up to 13 nm of diameter. We will show that this structure is preserved even in excess aqueous solution, where they form sponge-like nanoparticles (L3 NPs) in which two enzymes of different sizes, Aspartic protease and beta-galactosidase (34 KDa and 460 KDa, respectively), could be included. To reveal the nature of the interaction between the enzymes and the lipid matrix, we studied the adsorption of both proteins on the lipid layers formed by the L3 NPs. The results will be discussed in terms of the ability of these nanoparticles to encapsulate and release of the proteins in the lipid matrix.
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| 14. |
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| 15. |
- Bano, Fouzia, et al.
(författare)
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Single-molecule unbinding forces between the polysaccharide hyaluronan and its binding proteins
- 2018
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Ingår i: Biophysical Journal. - : Biophysical society. - 0006-3495 .- 1542-0086. ; 114:12, s. 2910-2922
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Tidskriftsartikel (refereegranskat)abstract
- The extracellular polysaccharide hyaluronan (HA) is ubiquitous in all vertebrate tissues, where its various functions are encoded in the supramolecular complexes and matrices that it forms with HA-binding proteins (hyaladherins). In tissues, these supramolecular architectures are frequently subjected to mechanical stress, yet how this affects the intermolecular bonding is largely unknown. Here, we used a recently developed single-molecule force spectroscopy platform to analyze and compare the mechanical strength of bonds between HA and a panel of hyaladherins from the Link module superfamily, namely the complex of the proteoglycan aggrecan and cartilage link protein, the proteoglycan versican, the inflammation-associated protein TSG-6, the HA receptor for endocytosis (stabilin-2/HARE), and the HA receptor CD44. We find that the resistance to tensile stress for these hyaladherins correlates with the size of the HA-binding domain. The lowest mean rupture forces are observed for members of the type A subgroup (i.e., with the shortest HA-binding domains; TSG-6 and HARE). In contrast, the mechanical stability of the bond formed by aggrecan in complex with cartilage link protein (two members of the type C subgroup, i.e., with the longest HA-binding domains) and HA is equal or even superior to the high affinity streptavidin⋅biotin bond. Implications for the molecular mechanism of unbinding of HA⋅hyaladherin bonds under force are discussed, which underpin the mechanical properties of HA⋅hyaladherin complexes and HA-rich extracellular matrices.
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| 16. |
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| 17. |
- Bengtsson, Elina, et al.
(författare)
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Altered Structural State of Actin Filaments Upon MYOSIN II Binding
- 2015
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Ingår i: Biophysical Journal. - : Biophysical Society. - 0006-3495 .- 1542-0086. ; 108:2 Suppl. 1, s. 299A-300A
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- The paths of actin filaments propelled over a heavy meromyosin (HMM) surface in the in vitro motility assay (IVMA) can statistically be described by a path persistence length (LPP) and has been hypothesized to be proportional to the flexural rigidity of the filaments. Here, we have studied the LPP at high (130 mM) ionic strength along with the persistence length of actin filaments in solution (LPS) to elucidate how HMM binding affects the flexural rigidity of actin filaments. Characterization and control of material properties, such as the path persistence length, is useful in engineered devices that takes advantages of the function of the muscle contractile proteins e.g. for biocomputation. It has been suggested that myosin binding reduces Lpp for phalloidin stabilizedact in filaments. This is consistent with the results presented here where the phalloidin stabilized actin filaments rigidity is reduced to the level of phalloidin free actin filaments in the IVMA. Further, reducing the MgATP concentration in the IVMA would increase the HMM head density along the actin filament hence making the effect of myosin binding more pronounced. A reduced [MgATP] from 1 mM to 0.02-0.05 mM did indeed reduce the LPP from 10-12 mm to 6-7 mm for both phalloidin-stabilized and phalloidin free actin filaments. Additionally, we found a negative correlation between the LPS and the [HMM]/actin ratio. However, this [HMM] dependent reduction observed in LPS was too small to account for the reduction in LPP seen with reduced [MgATP] in the IVMA. Monte-Carlo simulations and theoretical analysis revealed that the large reduction in LPP is consistent with the idea that every head attachment adds an extra angular displacement.(Support from EU-FP7-FET-ABACUS grant number 613044).
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| 18. |
- Bengtsson, Elina, et al.
(författare)
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Myosin-Induced Gliding Patterns at Varied [MgATP] Unveil a Dynamic Actin Filament
- 2016
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Ingår i: Biophysical Journal. - : Elsevier BV. - 0006-3495 .- 1542-0086. ; 111:7, s. 1465-1477
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Tidskriftsartikel (refereegranskat)abstract
- Actin filaments have key roles in cell motility but are generally claimed to be passive interaction partners in actin-myosin -based motion generation. Here, we present evidence against this static view based on an altered myosin-induced actin filament gliding pattern in an in vitro motility assay at varied [MgATP]. The statistics that characterize the degree of meandering of the actin filament paths suggest that for [MgATP] >= 0.25 mM, the flexural rigidity of heavy meromyosin (HMM)-propelled actin filaments is similar (without phalloidin) or slightly lower (with phalloidin) than that of HMM-free filaments observed in solution without surface tethering. When [MgATP] was reduced to <= 0.1 mM, the actin filament paths in the in vitro motility assay became appreciably more winding in both the presence and absence of phalloidin. This effect of lowered [MgATP] was qualitatively different from that seen when HMM was mixed with ATP-insensitive, N-ethylmaleimide-treated HMM (NEM-HMM; 25-30%). In particular, the addition of NEM-HMM increased a non-Gaussian tail in the path curvature distribution as well as the number of events in which different parts of an actin filament followed different paths. These effects were the opposite of those observed with reduced [MgATP]. Theoretical modeling suggests a 30-40% lowered flexural rigidity of the actin filaments at [MgATP] <= 0.1 mM and local bending of the filament front upon each myosin head attachment. Overall, the results fit with appreciable structural changes in the actin filament during actomyosin-based motion generation, and modulation of the actin filament mechanical properties by the dominating chemomechanical actomyosin state.
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| 19. |
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| 20. |
- Blau, Christian, et al.
(författare)
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Gromaps: A Gromacs-Based Toolset to Analyse Density Maps Derived from Molecular Dynamics Simulations
- 2019
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Ingår i: Biophysical Journal. - : Elsevier BV. - 0006-3495 .- 1542-0086. ; 116:1, s. 4-11
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Tidskriftsartikel (refereegranskat)abstract
- We introduce a computational toolset, named GROmaρs, to obtain and compare time-averaged density maps from molecular dynamics simulations. GROmaρs efficiently computes density maps by fast multi-Gaussian spreading of atomic densities onto a three-dimensional grid. It complements existing map-based tools by enabling spatial inspection of atomic average localization during the simulations. Most importantly, it allows the comparison between computed and reference maps (e.g., experimental) through calculation of difference maps and local and time-resolved global correlation. These comparison operations proved useful to quantitatively contrast perturbed and control simulation data sets and to examine how much biomolecular systems resemble both synthetic and experimental density maps. This was especially advantageous for multimolecule systems in which standard comparisons like RMSDs are difficult to compute. In addition, GROmaρs incorporates absolute and relative spatial free-energy estimates to provide an energetic picture of atomistic localization. This is an open-source GROMACS-based toolset, thus allowing for static or dynamic selection of atoms or even coarse-grained beads for the density calculation. Furthermore, masking of regions was implemented to speed up calculations and to facilitate the comparison with experimental maps. Beyond map comparison, GROmaρs provides a straightforward method to detect solvent cavities and average charge distribution in biomolecular systems. We employed all these functionalities to inspect the localization of lipid and water molecules in aquaporin systems, the binding of cholesterol to the G protein coupled chemokine receptor type 4, and the identification of permeation pathways through the dermicidin antimicrobial channel. Based on these examples, we anticipate a high applicability of GROmaρs for the analysis of molecular dynamics simulations and their comparison with experimentally determined densities.
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| 21. |
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| 31. |
- Czapla-Masztafiak, Joanna, et al.
(författare)
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Investigating DNA Radiation Damage Using X-Ray Absorption Spectroscopy
- 2016
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Ingår i: Biophysical Journal. - : Elsevier BV. - 0006-3495 .- 1542-0086. ; 110:6, s. 1304-1311
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Tidskriftsartikel (refereegranskat)abstract
- The biological influence of radiation on living matter has been studied for years; however, several questions about the detailed mechanism of radiation damage formation remain largely unanswered. Among all biomolecules exposed to radiation, DNA plays an important role because any damage to its molecular structure can affect the whole cell and may lead to chromosomal rearrangements resulting in genomic instability or cell death. To identify and characterize damage induced in the DNA sugar-phosphate backbone, in this work we performed x-ray absorption spectroscopy at the P K-edge on DNA irradiated with either UVA light or protons. By combining the experimental results with theoretical calculations, we were able to establish the types and relative ratio of lesions produced by both UVA and protons around the phosphorus atoms in DNA.
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| 32. |
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| 33. |
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| 34. |
- Dingeldein, Artur Peter Günther, et al.
(författare)
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Apoptotic Bax at Oxidatively Stressed Mitochondrial Membranes : Lipid Dynamics and Permeabilization
- 2017
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Ingår i: Biophysical Journal. - : Elsevier BV. - 0006-3495 .- 1542-0086. ; 112:10, s. 2147-2158
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Tidskriftsartikel (refereegranskat)abstract
- Mitochondria are crucial compartments of eukaryotic cells because they function as the cellular power plant and play a central role in the early stages of programmed cell death (apoptosis). To avoid undesired cell death, this apoptotic pathway is tightly regulated by members of the Bcl-2 protein family, which interact on the external surface of the mitochondria, i.e., the mitochondrial outer membrane (MOM), and modulate its permeability to apoptotic factors, controlling their release into the cytosol. A growing body of evidence suggests that the MOM lipids play active roles in this permeabilization process. In particular, oxidized phospholipids (OxPls) formed under intracellular stress seem to directly induce apoptotic activity at the MOM. Here we show that the process of MOM pore formation is sensitive to the type of OxPls species that are generated. We created MOM-mimicking liposome systems, which resemble the cellular situation before apoptosis and upon triggering of oxidative stress conditions. These vesicles were studied using P-31 solid-state magic-angle-spinning nuclear magnetic resonance spectroscopy and differential scanning calorimetry, together with dye leakage assays. Direct polarization and cross-polarization nuclear magnetic resonance experiments enabled us to probe the heterogeneity of these membranes and their associated molecular dynamics. The addition of apoptotic Bax protein to OxPls-containing vesicles drastically changed the membranes' dynamic behavior, almost completely negating the previously observed effect of temperature on the lipids' molecular dynamics and inducing an ordering effect that led to more cooperative membrane melting. Our results support the hypothesis that the mitochondrion-specific lipid cardiolipin functions as a first contact site for Bax during its translocation to the MOM in the onset of apoptosis. In addition, dye leakage assays revealed that different OxPls species in the MOM-mimicking vesicles can have opposing effects on Bax pore formation.
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| 35. |
- Dingeldein, Artur P G, et al.
(författare)
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BCL-2 Family Proteins Effect on Mitochondrial-Mimicking Membrane Structure by Solid State NMR
- 2015
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Ingår i: Biophysical Journal. - : Cell Press. - 0006-3495 .- 1542-0086. ; 108:2, s. 251A-252A
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- Mitochondria are not only the cells' powerhouse, but also involved in their suicide via apoptosis. Key regulators of this pathway are members of the Bcl-2 protein family which interact with the outer mitochondrial membrane to modulate permeability and enable the release of apoptotic stimuli like cytochrome c. For a long time the mitochondrial membrane forming lipids have been seen as merely structural building units with proteins doing the actual work. This view changed in recent years, since lipids were shown to be also directly involved in apoptotic events e.g. under intracellular oxidative stress. Oxidized phospholipids (OxPls) generated under these stress conditions might trigger mitochondria-mediated apoptosis. Their presence in mitochondrial membranes can severely alter the properties of these membranes with yet unknown consequences regarding the formation of pores through membrane-mediated interplay with apoptotic Bax protein. We therefore devised a model system that embodies oxidative stress conditions by incorporating OxPls into mitochondria mimicking model membranes composed of phosphatidylcholine (PC), phosphatidylethanolamine (PE) and cardiolipin (CL) to study the impact of OxPls on apoptotic Bax-membrane interactions. To obtain molecular insight into hydrophobic fatty acid regions of membranes and their hydrophilic interface which is responsible for first protein-membrane contacts, we used differential scanning calorimetry (DSC) and solid state NMR spectroscopy. Upon incorporating OxPls with carboxyl (PoxnoPC) or aldehyde (PazePC) groups at their truncated sn-2-chains into our mitochondria model membranes, calorimetric and NMR measurements showed dramatic changes. 31P NMR experiments revealed major perturbation effects in these membranes; an effect which presumably elevates the membrane binding of apoptotic Bax to the charged membranes and its partial penetration, being a prerequisite for its final formation of pores which enable cytochrome c release from the mitochondrial interior. Currently structural studies of various Bax-lipid assemblies are ongoing.
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| 36. |
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| 37. |
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| 38. |
- Elinder, Fredrik, et al.
(författare)
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Extracellular Linkers Completely Transplant the Voltage Dependence from Kv1.2 Ion Channels to Kv2.1
- 2016
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Ingår i: Biophysical Journal. - : CELL PRESS. - 0006-3495 .- 1542-0086. ; 111:8, s. 1679-1691
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Tidskriftsartikel (refereegranskat)abstract
- The transmembrane voltage needed to open different voltage-gated K (Kv) channels differs by up to 50 mV from each other. In this study we test the hypothesis that the channels voltage dependences to a large extent are set by charged amino-acid residues of the extracellular linkers of the Kv channels, which electrostatically affect the charged amino-acid residues of the voltage sensor S4. Extracellular cations shift the conductance-versus-voltage curve, G(V), by interfering with these extracellular charges. We have explored these issues by analyzing the effects of the divalent strontium ion (Sr2+) on the voltage dependence of the G(V) curves of wild-type and chimeric Kv channels expressed in Xenopus oocytes, using the voltage-clamp technique. Out of seven Kv channels, Kv1.2 was found to be most sensitive to Sr2+ (50 mM shifted G(V) by +21.7 mV), and Kv2.1 to be the least sensitive (+7.8 mV). Experiments on 25 chimeras, constructed from Kv1.2 and Kv2.1, showed that the large Sr2+-induced G(V) shift of Kv1.2 can be transferred to Kv2.1 by exchanging the extracellular linker between S3 and S4 (L3/4) in combination with either the extracellular linker between S5 and the pore (L5/P) or that between the pore and S6 (LP/6). The effects of the linker substitutions were nonadditive, suggesting specific structural interactions. The free energy of these interactions was similar to 20 kJ/mol, suggesting involvement of hydrophobic interactions and/or hydrogen bonds. Using principles from double-layer theory we derived an approximate linear equation (relating the voltage shifts to altered ionic strength), which proved to well match experimental data, suggesting that Sr2+ acts on these channels mainly by screening surface charges. Taken together, these results highlight the extracellular surface potential at the voltage sensor as an important determinant of the channels voltage dependence, making the extracellular linkers essential targets for evolutionary selection.
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| 39. |
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| 43. |
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| 44. |
- Gómez-Llobregat, Jordi, et al.
(författare)
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Anisotropic Membrane Curvature Sensing by Amphipathic Peptides
- 2016
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Ingår i: Biophysical Journal. - : Elsevier BV. - 0006-3495 .- 1542-0086. ; 110:1, s. 197-204
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Tidskriftsartikel (refereegranskat)abstract
- Many proteins and peptides have an intrinsic capacity to sense and induce membrane curvature, and play crucial roles for organizing and remodeling cell membranes. However, the molecular driving forces behind these processes are not well understood. Here, we describe an approach to study curvature sensing by simulating the interactions of single molecules with a buckled lipid bilayer. We analyze three amphipathic antimicrobial peptides, a class of membrane-associated molecules that specifically target and destabilize bacterial membranes, and find qualitatively different sensing characteristics that would be difficult to resolve with other methods. Our findings provide evidence for direction-dependent curvature sensing mechanisms in amphipathic peptides and challenge existing theories of hydrophobic insertion. The buckling approach is generally applicable to a wide range of curvature-sensing molecules, and our results provide strong motivation to develop new experimental methods to track position and orientation of membrane proteins.
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| 45. |
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| 46. |
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| 47. |
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| 48. |
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| 49. |
- Grønberg, Christina, et al.
(författare)
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Membrane Anchoring and Ion-Entry Dynamics in P-type ATPase Copper Transport
- 2016
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Ingår i: Biophysical Journal. - : Elsevier. - 0006-3495 .- 1542-0086. ; 111:11, s. 2417-2429
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Tidskriftsartikel (refereegranskat)abstract
- Cu+-specific P-type ATPase membrane protein transporters regulate cellular copper levels. The lack of crystal structures in Cu+-binding states has limited our understanding of how ion entry and binding are achieved. Here, we characterize the molecular basis of Cu+ entry using molecular-dynamics simulations, structural modeling, and in vitro and in vivo functional assays. Protein structural rearrangements resulting in the exposure of positive charges to bulk solvent rather than to lipid phosphates indicate a direct molecular role of the putative docking platform in Cu+ delivery. Mutational analyses and simulations in the presence and absence of Cu+ predict that the ion-entry path involves two ion-binding sites: one transient Met148-Cys382 site and one intramembranous site formed by trigonal coordination to Cys384, Asn689, and Met717. The results reconcile earlier biochemical and x-ray absorption data and provide a molecular understanding of ion entry in Cu+-transporting P-type ATPases.
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| 50. |
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