Effects of overlapping GB virus C/hepatitis G virus synthetic peptides on biomembrane models

The present study was undertaken to examine the physicochemical properties of three overlapping peptides belonging to the E2 envelope protein of Hepatitis G virus (GBV-C/HGV) and its interaction with phospholipid biomembrane models using biophysical techniques. We describe our findings concerning the surface activity and the interaction of the peptides with monolayers and liposomes composed of the zwitterionic phospholipids dipalmitoylphosphatidylcholine and dimyristoylphosphatidylcholine (DMPC) and a mixture of DMPC with the anionic phospholipid dimyristoylphosphatidylglycerol. The results inform about the effect of the chain length on their interaction with biomembrane models. The longest chain peptide interacts in a higher extent with all the phospholipid studied as a result of a combination of hydrophobic and electrostatic forces.     

Interaction of E2 and NS3 synthetic peptides of GB virus C/hepatitis G virus with model lipid membranes

The membrane-interacting properties of two potential epitopes of the GB virus C/Hepatitis G virus, located respectively at the regions (99-118) of the E2 structural protein and (440-460) of the NS3 non-structural protein were studied. Changes in the intrinsic fluorescence of Trp and Tyr residues after the addition of DPPC-LUV revealed that the peptide-membrane interaction was optimal above the gel-liquid crystalline transition temperature of the lipid. Differential scanning calorimetry studies showed that the E2 peptide incorporated into lipid bilayers perturbs the packing of lipids and affects their thermotropic properties. Moreover, the 20-mer structural peptide induced a slow leakage of vesicular contents at 55 degrees C.     

Interfacial properties of a synthetic peptide derived from hepatitis G virus E2 protein: interaction with lipid monolayers

A useful approach to get information about the potential fusogenic ability of virus synthetic peptides is the study of its interfacial properties and subsequent study in mono- and bilayers. In this work, we have characterized by means of physicochemical tools (i.e. compression isotherms and surface activity) the sequence 267-284, LLGTEVSEVLGGAGLTGG, derived from the E2 structural protein of HGV/GBV-C. The adsorption of the peptide at the air/water interface was monitored by following the increase in surface pressure as a function of time at two different pH values: 5 and 7. Parameters such as surface excess or molecular area were calculated from the equation of Gibbs. The peptide showed a tendency to migrate to the surface of a saline-buffered solution. It formed stable monolayers at the air/water interface giving a compression isotherm with a shape consistent with that of some alpha-helical peptide conformations. Brewster angle microscopy (BAM) showed that through compression the peptide formed multilayers. The studies with lipid monolayers (DPMC, DMPC/DMPG, and DMPC/DMTAP) showed that the peptide interacts with all the lipids assayed producing a marked disrupting effect upon them. In these effects electrostatic interactions seem to have some participation.     

Immobilized metal ion affinity chromatography of synthetic peptides. Binding via the alpha-amino group.

Peptides synthesized by the solid-phase method can be efficiently purified in a single immobilized metal affinity chromatography step based on interaction with the alpha-amino group if, after coupling of each amino acid residue, unreacted amino groups are irreversibly blocked by acetylation and if no strongly metal-binding amino acids (His, Trp, Cys) are present in the sequence. A difference in basicity for alpha- and epsilon-amino functions of ca. 2 pH units is sufficiently large to allow selective binding of peptides to immobilized metal ions via the unprotonated alpha-amino group. The binding is pH-dependent: on Cu(2+)- and Ni(2+)-loaded supports most peptides are maximally retarded at pH values around 7.5 and 8.5, respectively. The decreased binding strength at lower pH values is due to protonation of the alpha-amino function, whereas the reduced affinity at higher pH is caused by metal ion transfer from the matrix to the peptide. The metal ion is captured in a multidentate chelate where, in addition to the alpha-amino group, up to three adjacent deprotonated amide nitrogens are coordinated to the metal. If the pH is raised further, additional metal ions may be bound in biuret-like structures. Immobilized Ni2+, owing to its higher selectivity and affinity, is the preferred chromatographic support if slightly basic conditions can be tolerated.     

Mixed micelles containing sodium oleate: the effect of the chain length and the polar head group

We have investigated the mixing behavior of binary mixtures of the alkylglucosides (CnG) octyl beta-D-glucoside and decyl D-glucoside in combination with sodium oleate (NaOl), and the amine oxide surfactants (AO) N,N-dimethyldodecylamine oxide, N,N-bis (2-hydroxyethyl)dodecylamine oxide, and 3-lauramidopropyl-N,N-dimethylamine oxide in combination with NaOl. From the equilibrium surface tension measurements, the critical micelle concentration (cmc) data were obtained as functions of the composition. Values of the cmc were analyzed according to both the regular solution model developed by Rubingh for mixed micelles and Maeda's formulation for ionic/nonionic mixed micelles. Two interaction parameters, beta and B1, were estimated from the regular solution model and Maeda's formulation, respectively. For NaOl/CnG mixed systems, a decrease in the hydrocarbon chain length of CnG resulted in a stronger interaction with NaOl from both beta and B1 values. For NaOl/AO mixed systems, the bulkiness of a polar head group of AO surfactants influenced the interaction between NaOl and AO. The dynamic surface tension measurements show that all surface tension values of surfactant solutions examined decreased with the time. We found that the time dependence of surface tension values for NaOl mixed systems was greatly influenced by the presence of NaOl rather than the other component.     

Mechanisms for the selective gas-phase fragmentation reactions of methionine side chain fixed charge sulfonium ion containing peptides

To enable the development of improved tandem mass spectrometry based methods for selective proteome analysis, the mechanisms, product ion structures, and other factors influencing the gas-phase fragmentation reactions of methionine side-chain derivatized "fixed-charge" phenacylsulfonium ion containing peptide ions have been examined. Dissociation of these peptide ions results in the exclusive characteristic loss of the derivatized side chain, thereby enabling their selective identification. The resultant product ion(s) are then subjected to further dissociation to obtain sequence information for subsequent protein identification. Molecular orbital calculations (at the B3LYP/6-31 + G (d,p) level of theory) performed on a simple peptide model, together with experimental evidence obtained by multistage dissociation of a regioselectively deuterated methionine derivatized sulfonium ion containing tryptic peptide, indicate that fragmentation of the fixed charge containing peptide ions occurs via SN2 reactions involving the N- and C-terminal amide bonds adjacent to the methionine side chain, resulting in the formation of stable cyclic five- and six-membered iminohydrofuran and oxazine product ions, respectively. These studies further indicate that the rings formed via these neighboring group reactions are stable to further dissociation by MS3. As a consequence, the formation of b- or y-type sequence ions are "skipped" at the site of cyclization. Despite this, complete sequence information is still obtained because of the presence of both cyclic products.     

GB virus C (GBV-C) / hepatitis G virus (HGV): towards the design of synthetic peptides-based biosensors for immunodiagnosis of GBV-C/HGV infection

In the present study, new putative epitopes located in structural (E2) and non-structural (NS3) proteins of GBV-C/HGV were identified by computer-aided prediction of antigenicity and synthesized in solid-phase, following an Fmoc/tBut strategy, for their use in immunoassays. The corresponding synthetic peptides were used as antigens in ELISA assays and in real-time biospecific interaction measurements. This last approach allowed direct detection of GBV-C/HGV-specific antibodies in human sera. Good correlations were obtained between the biospecific interaction analysis and the ELISA. To verify the performance of these new assays in comparison to the existing recombinant E2 protein commercial test, antibodies to synthetic peptides were searched for in different panels of serum samples. The main conclusion of this work is the usefulness of E2 peptides in the detection of antibodies. Moreover, the NS3 peptide could be exploited to improve the sensitivity of the currently available test. Our results offer a new approach to develop new diagnostic peptide based biosensors for serodiagnosis of GBV-C/HGV infection.     

Thermotropic behavior of asymmetric chain length catanionic surfactants: the influence of the polar head group

Catanionic surfactants formed by the pairing of two ionic amphiphilic chains of opposite charge are now recognized as an important class of amphiphiles. Many aspects of their phase behavior have yet to be explored. In this work, two homologous series of catanionic surfactants were synthesized, based on the cationic headgroups trimethylammonium and pyridinium. Within each series, the headgroup and chain length of the cationic counterpart remains constant while for the anionic counterpart the headgroup is varied, while its alkyl chain length is also kept constant. Thus, one can directly monitor the influence of headgroup chemistry on the thermal behavior of these compounds. Differential scanning calorimetry (DSC) and polarizing light microscopy show that these compounds bear a rich and often complex thermotropic behavior, with the headgroup chemistry in some instances having a rather dramatic influence on phase behavior. Several liquid crystalline phases appear between the solid crystalline phase and the isotropic liquid phase. A qualitative correlation between the observed thermotropic behavior and the chemical nature of headgroup is presented.     

Effects of the phosphatidylglycerol head group on the binding of short dermcidin-derived peptides to the phospholipid membrane surface

Dermcidin (DCD) is a human peptide composed of 110 amino acids. When secreted into sweat, DCD undergoes postsecretory proteolytic processing to give the short antimicrobial peptides SSL-23 and SSL-25. As an initial phase of studies directed toward understanding the structural basis of the biological functions of these peptides, we chemically synthesized naturally occurring SSL-23 and SSL-25, as well as the artificial sequences SSL-21 and SSL-27, and analyzed their molecular interaction with bacterial and mammalian model surfaces. While dynamic-coating HPLC and CD spectroscopy revealed that the four SSL peptides selectively bound to a bacterial model membrane containing 1,2-dimyristoyl phosphatidylglycerol (DMPG) and underwent large structural changes, ³¹P NMR studies of the liposomes suggested that the attractive interaction between the peptides and DMPG did not lead to ion-pore formation or disruption of the model membrane. Our results strongly indicate that the SSL peptides express their selectivity to microorganisms by recognizing the head groups of their cell surface lipid.     

新型二胺扩链剂对合成聚脲机械性能的影响

以咔唑为原料合成了新型扩链剂N-乙基-3,6-二氨基咔唑;将其与端氨基聚醚和甲苯-2,4-二异氰酸酯(TDI)经溶液聚合合成了新型聚脲;采用红外光谱表征了新型扩链剂和聚脲的结构.结果表明,以N-乙基-3,6-二氨基咔唑作为扩链剂合成聚脲时,随异氰酸酯质量分数的增加,凝胶时间逐渐变短;所制备的聚脲的拉伸强度达25MPa,断裂伸长率降低为105%.     

Influence of charge state and amino acid composition on hydrogen transfer in electron capture dissociation of peptides

Although conventional N-Cα bond cleavage in electron capture dissociation (ECD) of multiply-charged peptides generates a complementary c′ and z′ fragment pair, the N-Cα cleavage followed by hydrogen transfer from c′ to z′ fragments produces other fragments, namely c′ and z′. In this study, the influence of charge state and amino acid composition on hydrogen transfer in ECD is described using sets of peptides. Hydrogen transferred ionic species such as c′ and z′ were observed in ECD spectra of doubly-protonated peptides, while the triply-protonated form did not demonstrate hydrogen transfer. The extent of hydrogen transfer in ECD of doubly-protonated peptides was dependent on constituent amino acids. The ECD of doubly-protonated peptides possessing numerous basic sites showed extensive hydrogen transfer compared with ECD of less basic peptides. The extent of hydrogen transfer is discussed from the viewpoints of the structure of peptide ions, the possibility of internal hydrogen bonding and intermediate lifetime of complex [c′+z′].     

Mixed-mode hydrophilic interaction/cation-exchange chromatography: separation of complex mixtures of peptides of varying charge and hydrophobicity

Mixed-mode hydrophilic interaction/cation-exchange chromatography (HILIC/CEX) was applied to the separation of two mixtures of synthetic peptide standards: (i) a 27-peptide mixture containing three groups of peptides (each group containing nine peptides of the same net charge of +1, +2 or +3), where the hydrophilicity/hydrophobicity of adjacent peptides within the groups varied only subtly (generally by only a single carbon atom); and (ii) peptide pairs with the same composition but different sequences, where the sole difference between the peptides was the position of a single amino acid substitution. HILIC/CEX is essentially CEX chromatography in the presence of high levels of organic modifier (generally ACN). The present study demonstrated the dramatic effect of increasing ACN concentration (optimum levels of 60-80%, depending on the application) on the separation of both mixtures of peptides. The greater the charge on the peptides, the better the separation achievable by HILIC/CEX. In addition, HILIC/CEX separation of both the peptide mixtures used in the present study was shown to be superior to that of the more commonly applied RP-HPLC mode. Our results highlight again the efficacy of HILIC/CEX as a peptide separation mode in its own right as well as an excellent complement to RP-HPLC.     

A novel, base-labile fluorous amine protecting group: synthesis and use as a tag in the purification of synthetic peptides

A new, base-labile fluorous tag based on the Msc amine protecting group was synthesized. Its use in the purification of synthetic peptides by fluorous HPLC or fluorous SPE was demonstrated.     

Influence of the interaction of chromophores of bis-cyanine dyes on the photogeneration of charge carriers in poly-N-epoxypropylcarbazole films

An increase in photoconductivity and in the extinguishing effect of an external electric field has been observed in the photoluminescence of doped poly-N-epoxypropylcarbazole films on changing from cyanine dyes with normal chromophores to the corresponding bis-cyanines. It was concluded that an increase in photogeneration of triplet electron-hole pairs arose with such replacement of the dye and that dissociation of the pairs was responsible for the photoconductivity. Translated from Teoreticheskaya i éksperimental'naya Khimiya, Vol. 34, No. 6, pp. 371–375, November–December, 1998.     

Effect of head group polarity and spacer chain length on the aggregation properties of gemini surfactants in an aquatic environment

The aggregation behavior of cationic gemini surfactants with respect to variation in head group polarity and spacer length is studied through conductance, surface tension, viscosity, and small-angle neutron-scattering (SANS) measurements. The critical micellar concentration (cmc), average degree of micelle ionization (beta(ave)), minimum area per molecule of surfactant at the air-water interface (A(min)), surface excess concentration (gamma(max)), and Gibb's free energy of micellization (delta G(mic)) of the surfactants were determined from conductance and surface tension data. The aggregation numbers (N), dimensions of micelles (b/a), effective fractional charge per monomer (alpha), and hydration of micelles (h(E)) were determined from SANS and viscosity data, respectively. The increasing head group polarity of gemini surfactant with spacer chain length of 4 methylene units promotes micellar growth, leading to a decrease in cmc, beta(ave), and delta G(mic) and an increase in N and b/a. This is well supported by the observed increase in hydration (h(E)) of micelles with increase in aggregation number (N) and dimension (b/a) of micelle.