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Effects of lipid composition and solution conditions on the mechanical properties of membrane vesicles.

Kato N, Ishijima A, Inaba T, Nomura F, Takeda S, Takiguchi K - Membranes (Basel) (2015)

Bottom Line: Liposomes prepared with a synthetic dimyristoylphosphatidylcholine, which has uniform hydrocarbon chains, were transformed easily compared with liposomes prepared using natural phosphatidylcholine.Surprisingly, bovine serum albumin or fetuin (soluble proteins that do not bind to membranes) decreased liposomal membrane rigidity, whereas the same concentration of sucrose showed no particular effect.These results show that the mechanical properties of liposomes depend on their lipid composition and environment.

View Article: PubMed Central - PubMed

Affiliation: Division of Biological Science, Graduate School of Science, Nagoya University, Chikusa-ku, Nagoya 464-8602, Japan. k614899x@m2.aichi-c.ed.jp.

ABSTRACT
The mechanical properties of cell-sized giant unilamellar liposomes were studied by manipulating polystyrene beads encapsulated within the liposomes using double-beam laser tweezers. Mechanical forces were applied to the liposomes from within by moving the beads away from each other, which caused the liposomes to elongate. Subsequently, a tubular membrane projection was generated in the tip at either end of the liposome, or the bead moved out from the laser trap. The force required for liposome transformation reached maximum strength just before formation of the projection or the moving out of the bead. By employing this manipulation system, we investigated the effects of membrane lipid compositions and environment solutions on the mechanical properties. With increasing content of acidic phospholipids, such as phosphatidylglycerol or phosphatidic acid, a larger strength of force was required for the liposome transformation. Liposomes prepared with a synthetic dimyristoylphosphatidylcholine, which has uniform hydrocarbon chains, were transformed easily compared with liposomes prepared using natural phosphatidylcholine. Surprisingly, bovine serum albumin or fetuin (soluble proteins that do not bind to membranes) decreased liposomal membrane rigidity, whereas the same concentration of sucrose showed no particular effect. These results show that the mechanical properties of liposomes depend on their lipid composition and environment.

No MeSH data available.


Related in: MedlinePlus

Cosedimentation assays of proteins with liposomes. β€˜βˆ’β€™, β€˜BSA’, and β€˜Fetuin’ indicate the protein assayed (β€˜βˆ’β€™ indicates that a solution containing liposomes made from PC and PG (1:1, mol/mol) was centrifuged in the absence of protein). β€˜M’, β€˜S’ and β€˜P’ indicate molecular weight markers, the supernatants and pellets of the centrifuged samples, respectively. All phospholipids used were obtained from native sources. HEPES-buffer alone or buffer containing 2.0 mg/mL BSA or 0.2 mg/mL fetuin was used for swelling the lipid films to prepare liposomes. The molecular weight of BSA is 66 kDa. It should be noted here that, even though the molecular weight of fetuin is about 50 kDa, this protein usually appears on the gel at the position of 50–70 kDa [47]. In the experimental conditions, if proteins bound to liposome membranes only weakly with a dissociation constant of the micro molar order, yet almost all proteins should be co-sedimentated to the precipitate fraction. However, only a very slight band of BSA and no band of Fetuin were detected in the precipitate fractions. Thus, we conclude that these proteins have little or no binding affinity to the membrane.
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membranes-05-00022-f005: Cosedimentation assays of proteins with liposomes. β€˜βˆ’β€™, β€˜BSA’, and β€˜Fetuin’ indicate the protein assayed (β€˜βˆ’β€™ indicates that a solution containing liposomes made from PC and PG (1:1, mol/mol) was centrifuged in the absence of protein). β€˜M’, β€˜S’ and β€˜P’ indicate molecular weight markers, the supernatants and pellets of the centrifuged samples, respectively. All phospholipids used were obtained from native sources. HEPES-buffer alone or buffer containing 2.0 mg/mL BSA or 0.2 mg/mL fetuin was used for swelling the lipid films to prepare liposomes. The molecular weight of BSA is 66 kDa. It should be noted here that, even though the molecular weight of fetuin is about 50 kDa, this protein usually appears on the gel at the position of 50–70 kDa [47]. In the experimental conditions, if proteins bound to liposome membranes only weakly with a dissociation constant of the micro molar order, yet almost all proteins should be co-sedimentated to the precipitate fraction. However, only a very slight band of BSA and no band of Fetuin were detected in the precipitate fractions. Thus, we conclude that these proteins have little or no binding affinity to the membrane.

Mentions: As proteins that do not bind to the lipid membrane, bovine serum albumin (BSA) and fetuin were used. We confirmed that BSA and fetuin do not bind to liposomes using a cosedimentation assay (Figure 5). BSA does not bind to liposomes, regardless of their lipid composition. Therefore, BSA is a protein that has been used as a control for cosedimentation assays between proteins and liposomes frequently used in our studies [17,21]. Fetuin, which is also obtained from bovine serum, is an acidic protein that is involved in the regulation of mineralization and bone formation [46]. Therefore, the electrostatic repulsion is one major reason why fetuin does not bind to liposomes containing acidic phospholipids, such as liposomes made from PC and PG.


Effects of lipid composition and solution conditions on the mechanical properties of membrane vesicles.

Kato N, Ishijima A, Inaba T, Nomura F, Takeda S, Takiguchi K - Membranes (Basel) (2015)

Cosedimentation assays of proteins with liposomes. β€˜βˆ’β€™, β€˜BSA’, and β€˜Fetuin’ indicate the protein assayed (β€˜βˆ’β€™ indicates that a solution containing liposomes made from PC and PG (1:1, mol/mol) was centrifuged in the absence of protein). β€˜M’, β€˜S’ and β€˜P’ indicate molecular weight markers, the supernatants and pellets of the centrifuged samples, respectively. All phospholipids used were obtained from native sources. HEPES-buffer alone or buffer containing 2.0 mg/mL BSA or 0.2 mg/mL fetuin was used for swelling the lipid films to prepare liposomes. The molecular weight of BSA is 66 kDa. It should be noted here that, even though the molecular weight of fetuin is about 50 kDa, this protein usually appears on the gel at the position of 50–70 kDa [47]. In the experimental conditions, if proteins bound to liposome membranes only weakly with a dissociation constant of the micro molar order, yet almost all proteins should be co-sedimentated to the precipitate fraction. However, only a very slight band of BSA and no band of Fetuin were detected in the precipitate fractions. Thus, we conclude that these proteins have little or no binding affinity to the membrane.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4384090&req=5

membranes-05-00022-f005: Cosedimentation assays of proteins with liposomes. β€˜βˆ’β€™, β€˜BSA’, and β€˜Fetuin’ indicate the protein assayed (β€˜βˆ’β€™ indicates that a solution containing liposomes made from PC and PG (1:1, mol/mol) was centrifuged in the absence of protein). β€˜M’, β€˜S’ and β€˜P’ indicate molecular weight markers, the supernatants and pellets of the centrifuged samples, respectively. All phospholipids used were obtained from native sources. HEPES-buffer alone or buffer containing 2.0 mg/mL BSA or 0.2 mg/mL fetuin was used for swelling the lipid films to prepare liposomes. The molecular weight of BSA is 66 kDa. It should be noted here that, even though the molecular weight of fetuin is about 50 kDa, this protein usually appears on the gel at the position of 50–70 kDa [47]. In the experimental conditions, if proteins bound to liposome membranes only weakly with a dissociation constant of the micro molar order, yet almost all proteins should be co-sedimentated to the precipitate fraction. However, only a very slight band of BSA and no band of Fetuin were detected in the precipitate fractions. Thus, we conclude that these proteins have little or no binding affinity to the membrane.
Mentions: As proteins that do not bind to the lipid membrane, bovine serum albumin (BSA) and fetuin were used. We confirmed that BSA and fetuin do not bind to liposomes using a cosedimentation assay (Figure 5). BSA does not bind to liposomes, regardless of their lipid composition. Therefore, BSA is a protein that has been used as a control for cosedimentation assays between proteins and liposomes frequently used in our studies [17,21]. Fetuin, which is also obtained from bovine serum, is an acidic protein that is involved in the regulation of mineralization and bone formation [46]. Therefore, the electrostatic repulsion is one major reason why fetuin does not bind to liposomes containing acidic phospholipids, such as liposomes made from PC and PG.

Bottom Line: Liposomes prepared with a synthetic dimyristoylphosphatidylcholine, which has uniform hydrocarbon chains, were transformed easily compared with liposomes prepared using natural phosphatidylcholine.Surprisingly, bovine serum albumin or fetuin (soluble proteins that do not bind to membranes) decreased liposomal membrane rigidity, whereas the same concentration of sucrose showed no particular effect.These results show that the mechanical properties of liposomes depend on their lipid composition and environment.

View Article: PubMed Central - PubMed

Affiliation: Division of Biological Science, Graduate School of Science, Nagoya University, Chikusa-ku, Nagoya 464-8602, Japan. k614899x@m2.aichi-c.ed.jp.

ABSTRACT
The mechanical properties of cell-sized giant unilamellar liposomes were studied by manipulating polystyrene beads encapsulated within the liposomes using double-beam laser tweezers. Mechanical forces were applied to the liposomes from within by moving the beads away from each other, which caused the liposomes to elongate. Subsequently, a tubular membrane projection was generated in the tip at either end of the liposome, or the bead moved out from the laser trap. The force required for liposome transformation reached maximum strength just before formation of the projection or the moving out of the bead. By employing this manipulation system, we investigated the effects of membrane lipid compositions and environment solutions on the mechanical properties. With increasing content of acidic phospholipids, such as phosphatidylglycerol or phosphatidic acid, a larger strength of force was required for the liposome transformation. Liposomes prepared with a synthetic dimyristoylphosphatidylcholine, which has uniform hydrocarbon chains, were transformed easily compared with liposomes prepared using natural phosphatidylcholine. Surprisingly, bovine serum albumin or fetuin (soluble proteins that do not bind to membranes) decreased liposomal membrane rigidity, whereas the same concentration of sucrose showed no particular effect. These results show that the mechanical properties of liposomes depend on their lipid composition and environment.

No MeSH data available.


Related in: MedlinePlus