Ryoichi Kuboi

Catechol derivatives inhibit the fibril formation of amyloid-β peptides

The inhibition of fibril formation of amyloid beta proteins (A beta) would be attractive therapeutic targets for the treatment of Alzheimers disease (AD). Dopamine (DA) and other catechol derivatives were used as inhibitory factors for A beta fibril formation. The fibril formation of A beta was monitored by Thioflavin T fluorescence, a transmission electron microscopy (TEM) and a total internal reflection fluorescence microscopy (TIRFM). Catechol and its derivatives showed the dose-dependent inhibitory effects on the spontaneous A beta fibril formation. The inhibitory activity depended on the chemical structure of catechol derivatives both in the presence and absence of the liposome a model of biomembrane. Formation of catechol quinone-conjugated-A beta adduct by a Schiff-base is a key step for the inhibition effect of A beta fibril formation.

Relationship between the mobility of phosphocholine headgroups of liposomes and the hydrophobicity at the membrane interface: A characterization with spectrophotometric measurements

In this study, we investigated the dynamics of a membrane interface of liposomes prepared by eight zwitterionic phosphatidylcholines in terms of their headgroup mobility, with spectroscopic methods such as dielectric dispersion analysis (DDA), fluorescence spectroscopy. The DDA measurement is based on the response of the permanent dipole moment to a driving electric field and could give the information on the axial rotational Brownian motion of a headgroup with the permanent dipole moment. This motion depended on kinds of phospholipids, the diameter of the liposomes, and the temperature. The activation energy required to overcome the intermolecular force between headgroups of phospholipids depended on the strength of the interaction between headgroups such as hydrogen bonds and/or dipole-dipole interaction. Hydration at the phosphorous group of phospholipid and the molecular order of lipid membrane impaired the interaction between headgroups. Furthermore, the hydrophobicity of membrane surface increased parallel to the increase in headgroup mobility. It is, therefore, concluded that hydration of headgroup promoted its mobility to make the membrane surface hydrophobic. The lipid membrane in liquid crystalline phase or the lipid membrane with the larger curvature was more hydrophobic.

Charged liposome affects the translation and folding steps of in vitro expression of green fluorescent protein

The role of the charged liposome on the in vitro expression of green fluorescent protein (GFP) was investigated, focusing on its elemental steps such as transcription, translation and folding. The total GFP expression was enhanced to 145% when a neutral liposome (POPC: 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocoline) was added externally to a cell-free translation system. On the contrary, the addition of the charged liposome composed of POPC with anionic 1-palmitoyl-2-oleoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (POPG) or cationic stearyl amine (SA) inhibited the total GFP expression, depending on the surface charge density of liposome. In transcription, the RNA synthesis was enhanced regardless of the variation of the surface charge, indicating that transcription was enhanced due to the stabilization of RNA structure by its hydrophobic interaction with liposome. Translation was inhibited by cationic liposome although it was enhanced by anionic liposome and neutral liposome. On the other hand, the folding was not inhibited in the presence of neutral liposome, whereas anionic liposome and cationic liposome inhibited the folding in proportion to the their surface charges, suggesting that the total GFP expression was controlled by a charged liposome in the translation step and folding step.

Detection of a heat stress-mediated interaction between protein and phospholipid membrane using dielectric measurement

The dielectric response of lipid bilayer membrane vesicles (liposomes) prepared using either phosphatidylcholine from egg (EPC) or 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) was analyzed at a frequency range of 0.1 to 100 MHz. A marked dielectric dispersion for EPC and POPC liposome suspensions was observed above 1 MHz. An appropriate analysis of the dielectric dispersion curve was performed using the Cole-Cole equation and the Debye equation and was found to provide a method for the determination of dielectric parameters. Among the dielectric parameters, the characteristic frequency of a second dispersion around 50 MHz varied corresponding with changes in the test conditions. Of particular note is that an anomalous change in the characteristic frequency in the presence of protein corresponded to the degree of hydrophobic interaction between proteins and liposomes. The value of the frequency around 50 MHz, as well as the decrease in permittivity over the frequency range tested, are indicators of the interaction between proteins and liposomes.

Immobilization of intact liposomes on solid surfaces: a quartz crystal microbalance study.

Immobilization of model cell membranes (liposomes) on sensing devices is useful for the detection, evaluation, and analysis of interaction between cell membranes and proteins or other biological materials. The method to immobilize the liposomes on the solid surface was investigated by using the quartz crystal microbalance (QCM) method, focusing on the density of immobilized liposomes and their stability. The different liposomes were applied onto the planar surface of the QCM electrode by varying the immobilization method. The results showed that the immobilized liposomes depended not only on the immobilization method but also on the properties of the liposomes. The liposome with low membrane fluidity likely showed intact immobilization on a solid surface. An electrostatic interaction also affected the amount of immobilized liposomes on the functionalized quartz crystal. By using the amino-coupling method, intact liposomes could be immobilized on a solid surface and immobilized liposomes could stabilize for 10 h with frequency changes less than 5%.

Liposomes destabilize tRNA during heat stress

Biomembranes play an important role in cellular response to heat stress. In this study, we focus on the interaction between liposomes and tRNA. Upon heat treatment we determined circular dichroism spectra of tRNA in presence of liposomes prepared from POPC (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine) and cholesterol (Ch). To compare thermal stability, midpoint temperature (T(m)) of tRNA was calculated from normalized theta(208). Addition of POPC/Ch liposomes decreased the T(m) value of tRNA from 48 degrees C to 38 degrees C. We conclude that POPC/Ch liposomes interact with tRNA and destabilize its conformation under heat stress.

Arsenic (V) induces a fluidization of algal cell and liposome membranes.

Arsenate is one of the most poisonous elements for living cells. When cells are exposed to arsenate, their life activities are immediately affected by various biochemical reactions, such as the binding of arsenic to membranes and the substitution of arsenic for phosphate or the choline head of phospholipids in the biological membranes. The effects of arsenate on the life activities of algae Chlorella vulgaris were investigated at various concentrations and exposure times. The results demonstrated that the living activities of algal cells (10(10)cells/L) were seriously affected by arsenate at a concentration of more than 7.5mg As/L within 24h. Algal cells and the artificial membranes (liposomes) were exposed to arsenate to evaluate its effects on the membrane fluidization. In the presence of arsenate, the membranes were fluidized due to the binding and substitution of arsenate groups for phosphates or the choline head on the their membrane surface. This fluidization of the biological membranes was considered to enhance the transport of toxicants across the membrane of algal cells.

Evaluation of interaction between liposome membranes induced by stimuli responsive polymer and protein

Immobilized liposome chromatography was utilized as a novel method for the quantitative evaluation of the interaction between liposome membranes. The capacity factors evaluated from the elution profile showed that interaction between 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) liposome membranes occurred in the presence of a stimuli responsive polymer and protein under specific stimulus conditions. The occurrence of such interaction was supported by experimental results for POPC liposome membrane fusion under corresponding stimuli conditions.

Inversion of the Configuration of a Single Stereocenter in a β-Heptapeptide Leads to Drastic Changes in its Interaction with Phospholipid Bilayers

One small change for a molecule The change in the configuration of a single amino acid in a β3-heptapeptide can lead to remarkable changes in the physicochemical properties of the peptide with respect to the interaction with lipid vesicles (liposomes) formed from DOPC (1,2-dioleoyl-sn-glycero-3-phosphocholine).

Copper-mediated growth of amyloid β fibrils in the presence of oxidized and negatively charged liposomes

Amyloid β protein (Aβ) from Alzheimers disease formed fibrillar aggregates and their morphology depended on oxidized and negatively charged liposomes. The morphology of fibrillar aggregates was affected by Cu(2+), together with their growth kinetics. This is because Cu(2+) inhibited the nucleation step in the formation of amyloid Aβ fibrillar aggregates by forming Aβ/Cu complex inactive to the growth of fibrillar aggregates. In addition, this is probably because Cu(2+) affected the fibrillar aggregate formed on the surface of liposomes. These findings would give a better understanding of the formation mechanism of amyloid fibrils on biomembranes.