Determination of available breaking stress of agar and gellan gum plate culture methods and the duration of bacterial culture under strong acidic conditions.

AIMS: Several acidophilic bacteria have not been cultured, primarily owing to the lack of suitable culture methods under strong acidic conditions. This study aimed to quantitatively evaluate the strengths of the agar plates (AP) and gellan gum plates (GP), and optimal culture periods under strong acidic conditions. METHODS AND RESULTS: To define the lower limit of plate strength for bacterial isolation culture, the diameter of Escherichia coli K12 colonies and the breaking stress of plates at different concentrations of gelling agents, medium composition and pH conditions were determined. The lower limit of available strength of AP and GP was 19·6 and 14·8 kPa, respectively. Medium composition slightly affected AP breaking stress, although GP with a high cationic concentration medium could not be prepared. CONCLUSIONS: Assessment of the strength limits of AP and GP revealed that AP is not suitable for prolonged bacterial culture (≥72 h). Furthermore, GP was completely ineffective for bacterial culture under highly acidic conditions (≤pH 1·0). SIGNIFICANCE AND IMPACT OF THE STUDY: Our quantitative evaluation method based on breaking stress is a potentially valuable tool to understand the state and the suitable limit of plate culture methods in more detail under various conditions.

Effect of tetrasodium pyrophosphate concentration and cooking time on the physicochemical properties of process cheese.

Tetrasodium pyrophosphate (TSPP) is widely used as an emulsifying salt (ES) in process cheese. Previous reports have indicated that TSPP exhibits some unusual properties, including the gelation of milk proteins at specific ES concentrations. We studied the effect of various concentrations (0.25-2.75%) of TSPP and cooking times (0-20min) on the rheological, textural, and physical properties of pasteurized process Cheddar cheese using a central composite rotatable experimental design. Cheeses were made with a constant pH value to avoid pH as a confounding factor. Modeling of the textural properties of process cheese made with TSPP exhibited complex behavior, with polynomial models (cubic) giving better predictions (higher coefficient of determination values) than simpler quadratic models. Meltability indices (degree of flow from the UW MeltProfiler (University of Wisconsin-Madison), loss tangent value at 60°C from rheological testing, and Schreiber melt area) initially decreased with increasing TSPP concentrations, but above a critical ES concentration (~1.0%) meltability increased at higher TSPP concentrations. The storage modulus values measured at 70°C for process cheese initially increased with increasing TSPP concentration, but above a concentration of 1% ES, the storage modulus values decreased. Cooking time had little effect on the various melting or rheological properties. With an increase in TSPP concentration, the insoluble Ca and P contents increased, suggesting that TSPP addition resulted in the formation of insoluble calcium pyrophosphate complexes; some of which were likely associated with caseins. A portion of the added TSPP remained in the soluble phase. The acid-base buffering profiles also indicated that calcium pyrophosphate complexes were formed in cheese made with TSPP. In milk systems, low levels of TSPP have been shown to induce protein crosslinking and gelation, whereas at higher TSPP concentrations milk gelation was inhibited due to excessive charge repulsion from these calcium pyrophosphate complexes. We hypothesized that a similar phenomenon was occurring in our process cheese, resulting in the initial reduction in meltability with TSPP addition due to protein crosslinking, but at higher TSPP levels meltability increased due to excessive charge repulsion.

RISING beamline (BL28XU) for rechargeable battery analysis.

The newly installed BL28XU beamline at SPring-8 is dedicated to in situ structural and electronic analysis of rechargeable batteries. It supports the time range (1 ms to 100 s) and spatial range (1 µm to 1 mm) needed for battery analysis. Electrochemical apparatus for battery charging and discharging are available in experimental hutches and in a preparation room. Battery analysis can be carried out efficiently and effectively using X-ray diffraction, X-ray absorption fine-structure analysis and hard X-ray photoelectron spectroscopy. Here, the design and performance of the beamline are described, and preliminary results are presented.

Impact of cystic fibrosis transmembrane conductance regulator gene mutation on the occurrence of chronic pancreatitis in Japanese patients.

DNA analyses of the cystic fibrosis transmembrane conductance regulator (CFTR) gene in Japanese patients with idiopathic chronic pancreatitis (ICP) were performed to determine the relationship between the CFTR mutation and ICP. The study included patients with alcoholic pancreatitis (n = 20), patients with ICP (n = 20) and healthy volunteers (controls; n = 110). The poly-T region in intron 8 of the CFTR gene was analysed by direct sequencing. The CFTR coding region was screened using single-strand conformational polymorphism and direct sequencing. In the controls, frequencies of the 5T genotype and 5T allele were 4.5% and 3.6%, respectively. The frequency of the 5T genotype was significantly higher in the ICP group (20%) versus controls, but was not significantly different in alcoholic chronic pancreatitis patients (5%). Thus, the CFTR gene mutation, especially the 5T genotype, appears to have some relationship to ICP prevalence in Japanese patients independent of cystic fibrosis.

Pulmonary exposure to carbon black nanoparticles increases the number of antigen-presenting cells in murine lung.

Particulate matters can enhance antigen-related airway inflammation and immunoglobulin production. The present study was designed to determine the effects of different sizes of nanoparticles on the antigen-presenting cells (APC) in the lung. ICR mice were exposed to vehicle, carbon black (CB) nanoparticles (14 nm or 56 nm), ovalbumin (OVA), or OVA + nanoparticles intratracheally. The expression of major histocompatibility complex (MHC) class II, costimulatory molecules (CD80, CD86, CD11c), and DEC205 (dendritic cell marker), F4/80 (macrophage marker), and CD19 (B-cell marker) in the lung cells was measured by flow cytometry. 14 nm nanoparticles, but not 56 nm nanoparticles, increased the number of the total lung cells. Combination of OVA and 14 nm or 56 nm nanoparticles increased the total lung cells. The expression of MHC class II and/or costimulatory molecules and the number of APC in the lung were increased by 14 nm nanoparticles in the presence or absence of OVA. The increases were more prominent with combination of OVA and 14 nm nanoparticles. 56 nm nanoparticles did not show any significant effects. 14 nm CB nanoparticles can increase the expression of MHC class II and costimulatory molecules and the number of APC in the lung, especially in the presence of antigen, which can result in subsequent antigen-related airway inflammation and immunoglobulin production.

Development of a novel artificial medium based on utilization of algal photosynthetic metabolites by symbiotic heterotrophs.

AIMS: (i) Quantitative and qualitative analyses of photosynthetic metabolites of Chlorella sorokiniana and elucidation of the mechanism of their utilization by algal symbionts. (ii) Development of artificial medium that imitates photoautotroph-heterotroph interaction and investigation of its suitability for isolation of novel microbes from the environment. METHODS AND RESULTS: Various components, including free dissolved carbohydrates, nitrogenous compounds and vitamin, were detected and together contributed 11.1% (as carbon content) of the total photosynthetic metabolites in the medium. Utilization of these photosynthetic metabolites in algal culture broth by algal symbionts was studied. Many symbionts showed specific utilization patterns. A novel artificial extracellular released organic carbon medium, which imitated the nutritional conditions surrounding algae, was developed based on the pattern of utilization of the algal metabolites by the symbiotic heterotrophs. About 42.9% of the isolates were closely related to photoautotrophic-dependent and oligotrophic bacteria. CONCLUSIONS: With the novel artificial medium, it was possible to selectively isolate some bacterial strains. SIGNIFICANT AND IMPACT OF THE STUDY: Synthetic bacterial growth medium is an important and basic tool for bacterial isolation from environmental samples. The current study shows that preferential separation of typical bacterial subset can be achieved by using artificial medium that mimics photosynthetic metabolites.

UV mutagenesis of Cupriavidus necator for extracellular production of (R)-3-hydroxybutyric acid.

AIM: Ultraviolet (UV) mutagenesis was carried out to obtain mutant strains of Cupriavidus necator that could produce (R)-3-hydroxybutyric acid [(R)-3-HB] in the culture supernatant. METHODS AND RESULTS: C. necator (formerly known as Ralstonia eutropha) was subjected to UV radiation to generate mutants that are capable of producing (R)-3-HB in the culture supernatant. Results indicated that UV mutagen disrupted the phbB (phbB knock-out) and thus, promoted production of (R)-3-HB in mutant strains. Inclusion of acetoacetate esters (carbonyl compounds) in the culture broth led to increased production of (R)-3-HB. Thus, acetoacetyl-CoA (an intermediate of the PHB synthetic pathway) might have been converted to acetoacetate, which in the presence of (R)-3-HB dehydrogenase and NADPH/NADP(+), resulted in extracellular production of (R)-3-HB. CONCLUSIONS: UV mutagenesis proved to be a satisfactory method in generating interesting mutants for extracellular production of (R)-3-HB. Extracellular production of (R)-3-HB upon addition of acetoacetate esters would suggest a likely (R)-3-HB biosynthetic pathway in C. necator. SIGNIFICANCE AND IMPACT OF THE STUDY: Mutants obtained in this study are very useful for production of (R)-3-HB. For the first time, the production of (R)-3-HB by C. necator via acetoacetate is reported.

Photobioreactors for mass cultivation of algae.

Algae have attracted much interest for production of foods, bioactive compounds and also for their usefulness in cleaning the environment. In order to grow and tap the potentials of algae, efficient photobioreactors are required. Although a good number of photobioreactors have been proposed, only a few of them can be practically used for mass production of algae. One of the major factors that limits their practical application in algal mass cultures is mass transfer. Thus, a thorough understanding of mass transfer rates in photobioreactors is necessary for efficient operation of mass algal cultures. In this review article, various photobioreactors that are very promising for mass production of algae are discussed.

Interactions of N-terminal fragments of groups I and II phospholipases A2 with phospholipid bilayers and their surface recognition properties.

In order to elucidate the roles of the N-terminal segments of groups I and II phospholipases A2 (PLA2s) which have been known to have alpha-helical structure and have been assumed to be involved in the water/lipid interface recognition site, the peptides corresponding to the N-terminal moieties of group I PLA2 (Naja naja atra) and group II PLA2s (Trimeresurus flavoviridis and Crotalus atrox) were synthesized and their interactions with model membranes were studied. Circular dichroism spectra showed that N-terminal peptides of both groups I and II PLA2s took alpha-helical structure in trifluoroethanol but no significant secondary structure in buffer (pH 8.0). In the presence of acidic liposomes, N-terminal fragments of group II PLA2s formed alpha-helical structure, while that of group I PLA2 remained unaffected. The hydrophobic moments showed that amphipathicities of N-terminal fragments of group II PLA2s are evidently larger than those of N-terminal fragments of group I PLA2s. The leakage of carboxyfluorescein from acidic liposomes was induced only with group II PLA2 peptides. Large blue shift and increase in intensity of tryptophan fluorescence were also observed for group II PLA2 peptides when interacting with acidic liposomes. Such difference in the modes of interactions with lipid bilayers between N-terminal peptides of groups I and II PLA2s appears to be due in large part to the difference in intrinsic alpha-helix forming properties of their amino acid sequences. It is inferred that N-terminal amphipathic alpha-helical structures of group I PLA2s are possibly formed by assistance of a neighboring chain bridged by Cys-11 and Cys-77.

Relationship between antimicrobial activity and amphiphilic property of basic model peptides.

Several cationic model peptides of the prepiece moieties of mitochondrial protein precursors were found to be active against Gram-positive bacteria, but inactive against Gram-negative bacteria. The CD spectra of the model peptides in the presence of phospholipid liposomes demonstrated that antimicrobial activity was generally in parallel with the content of the alpha-helical amphiphilicity. The results indicate that appropriate positioning of cationic and hydrophobic groups in the stable alpha-helical structure of the peptides is important to exhibit antimicrobial activity. These peptides also have an ability to leak carboxyfluorescein from acidic and neutral phospholipid vesicles, suggesting that the peptides interact with the bacterial membrane to perturb it.

Effect of amphipathic peptides with different alpha-helical contents on liposome-fusion.

The peptide-induced fusion of neutral and acidic liposomes was studied in relation to the amphiphilicities evaluated by alpha-helical contents of peptides by means of a carboxyfluorescein leakage assay, light scattering, a membrane intermixing assay and electron microscopy. An amphipathic mother peptide, Ac-(Leu-Ala-Arg-Leu)3-NHCH3 (4(3], and its derivatives, [Pro6]4(3) (1), [Pro2,6]4(3) (2), and [Pro2,6,10]4(3) (3), which have very similar hydrophobic moments, caused a leakage of contents from small unilamellar vesicles composed of egg yolk phosphatidylcholine and egg yolk phosphatidic acid (3:1). The abilities of the peptides to induce the fusion of the acidic liposomes increased with increasing alpha-helical content: in acidic liposomes the helical contents were in the order of 4(3) greater than 1 greater than 2 greater than 3 (Lee et al. (1989) Chem. Lett., 599-602). Electron microscopic data showed that 1 caused a transformation of the small unilamellar vesicles (20-50 nm in diameter) to large ones (100-300 nm). Based on the fact that these peptides have very similar hydrophobic moments despite of decreasing in the mean residue hydrophobicities to some extent, it was concluded that the abilities of the peptides to induce the fusion of liposomes depend on the extent of amphiphilic conformation evaluated by alpha-helical contents of the peptides in the presence of liposomes. For neutral liposomes of egg yolk phosphatidylcholine, all the proline-containing peptides showed no fusogenic ability but weak leakage abilities, suggesting that the charge interaction between the basic peptides and acidic phospholipid is an important factor to induce the perturbation and fusion of the bilayer.

Two mode ion channels induced by interaction of acidic amphipathic alpha-helical peptides with lipid bilayers.

In order to investigate the ion permeability and selectivity of ion channel formed by amphipathic alpha-helical peptides, we designed to synthesize an acidic peptide Ac(Leu-Ala-Glu-Leu)3NHCH3 (Glu-4(3)) and its channel property was compared with a basic peptides Arg-4(3) in which Glu in Glu-4(3) was replaced by Arg. Two modes of the conductance change were observed by the interaction of Glu-4(3) with planar lipid bilayers; a steady increase of the conductance with the elapse of time (mode 1) and a spike-like increase of the current (mode 2) appearing with a lag time and overlapping the model current increase. The application of negative membrane potential induced the mode 1 current and the lowering pH decreased it, suggesting that the mode 1 current is caused by slow insertion of Glu-4(3) into the lipid bilayer and then by forming certain unknown bundles like semichannels. Mode 2 was found to be consisted of channel type opened-close current with several different conductances and relatively short opening lifetimes. There was no ion selectivity in the mode 1 current, whereas the mode 2 current was cation selective. The peptide Arg-4(3) has formed a cation-selective ion channel but not shown such two mode current changes. The membrane potential formation experiment in liposomes using DiSC3(5) also showed the cation selectivity for Arg-4(3) and non-ion selectivity for Glu-4(3). The difference between Arg-4(3) and Glu-4(3) was also observed in conformational analysis by CD and in dye-release experiment from liposome. Such difference was discussed in terms of electrostatic interaction between peptides and lipid head groups.

Formation of ion channels in planar lipid bilayer membranes by synthetic basic peptides.

We made use of a planar lipid bilayer system to examine the action of synthetic basic peptides which model the prepiece moiety of mitochondrial protein precursors and have antibacterial activity against Gram-positive bacteria. The sequences of the peptides used were as follows: Ac-(Ala-Arg-Leu)3-NHCH3 (3(3], Ac-(Leu-Ala-Arg-Leu)2-NHCH3 (4(2], Ac-(Leu-Ala-Arg-Leu)3-NHCH3 (4(3], Ac-(Leu-Leu-Ala-Arg-Leu)2-NHCH3 (5(2]. These peptides interacted differently with planar lipid bilayer membranes and membrane conductance increased by the formation of ion channels. The effects of the peptides on the macroscopic current-increase and on the probability of channel formation, at the single channel level were in the order of 4(3) greater than 4(2) approximately 5(2) much greater than 3(3), a finding which correlates with the antibacterial activity of these peptides. The micromolar (microM) order concentration at which the channel was formed resembles that causing antibacterial activity. Thus, the peptide antibacterial activity may occur through an increase in ion permeability of the bacterial membrane. The single-channel properties were investigated in detail using 4(3), the peptide with the highest ion channel-forming activity. Many types of channels were observed with respect to conductance (2-750 pS) and voltage dependency of gating. However, the channels were all cation-selective. These results suggest that the ion channels formed by peptide 4(3) may be able to take on a variety of conformations and/or assembly.

Design and synthesis of basic peptides having amphipathic beta-structure and their interaction with phospholipid membranes.

Basic amphipathic beta-structural peptides, Ac-(Ser-Val-Lys-Val)n-NHCH3 (1n, n = 1-3) and Ac-(Lys-Val)n-NHCH3 (2n, n = 2-4), were synthesized and their interaction with DPPC and DPPC-DPPG (3:1) bilayers was studied by CD, dye-leakage and fluorescence experiments. The CD data indicated that oligopeptides consisting of more than eight residues with alternating hydrophobic (Val) and hydrophilic amino acids (Ser and Lys) were able to form an amphipathic beta-structure in acidic phospholipid bilayers, but not or weakly in aqueous solution and in neutral phospholipid bilayers. The dye-leakage experiment showed that the basic amphipathic beta-structural peptides interact with acidic phospholipid bilayers to perturb them, but less effectively compared with basic amphipathic alpha-helical peptides. Fluorescent spectroscopic data suggest that hydrophobic side of the amphipathic peptides may immerse into membrane without deep penetration. Based on these results, we postulate that the formation of the basic amphipathic beta-structure on acidic lipid bilayers may be due to the combined effect of electrostatic and hydrophobic interactions between basic peptides and acidic lipid bilayers.

The spectroscopic analysis for binding of amphipathic and antimicrobial model peptides containing pyrenylalanine and tryptophan to lipid bilayer.

The binding of basic amphipathic fluorescent peptides to lipid bilayers was studied in relation to their antimicrobial activity. Four fluorescent peptides containing pyrenylalanine or tryptophan in an amphipathic basic peptide (4(4] consisting of four repeated units of tetrapeptide, -L-Leu-L-Ala-L-Arg-L-Leu-, were found to have antimicrobial activities against Gram-positive bacteria and to take conformations with fairly high alpha-helical content both in aqueous solutions and liposomes. The fluorescence spectroscopic data suggested that the pyrenylalanine-peptide existed as a monomer in methanol or liposomes but as an oligomer in aqueous solutions to form an excimer between pyrenylalanyl residues. Upon binding with liposomes, the fluorescence spectra of the tryptophan-containing peptide shifted to a shorter wavelength, indicating the change in the state of tryptophan from hydrophilic environment to hydrophobic one. The analytical data for the quenching of tryptophan fluorescence by I- anion suggest that the tryptophan residue in the peptide is not deeply buried in the hydrophobic core of the bilayers. Based on these findings, it is suggested that the peptides may interact with liposomes in such a manner that they lie parallel to the surface of the lipid bilayers with their hydrophobic regions shallowly in the amphipathic moiety of the bilayers.

Effect of salts on conformational change of basic amphipathic peptides from beta-structure to alpha-helix in the presence of phospholipid liposomes and their channel-forming ability.

A synthetic model peptide, H-(Leu-Al alpha-Arg-Leu)3-(Leu-Arg-Al alpha-Leu)3-OH (4(6)) can form ion channels in planar lipid bilayers by taking an amphipathic alpha-helix (Agawa, Y., Lee, S., Ono, S., Aoyagi, H., Ohno, M., Taniguchi, T., Anzai, K. and Kirino, Y. (1991) J. Biol. Chem. 266, 20218-20222). For further study of ion channels formed by this type of peptides, we planned to synthesize [Trp1]-4(6)(Ser) and [Trp12]-4(6)(Ser) in which a hydrophilic amino acid, Ser, was introduced in several positions of 4(6) instead of hydrophobic ones. This modification was expected to decrease the ability of membrane perturbation and to simplify various current levels of the channel observed for 4(6). Furthermore, additional Trp was introduced to the N-terminus or position 12 to monitor the lipid-peptide interaction. CD study showed that both peptides formed a random structure in buffer, but an alpha-helix in the presence of egg PC and a beta-structure in egg PC/egg PG (3:1). Moreover, addition of NaCl to the acidic liposomes induced the conformational transition in the peptide from beta-structure to alpha-helix. Salt-induced conformational transition in the presence of acidic liposomes was discussed in terms of membrane binding and ion-channel formation in planar lipid bilayer. Despite introduction of hydrophilic residues instead of hydrophobic residues in 4(6), the peptide showed nearly the same dye-release ability from egg PC- egg PG liposomes as 4(6). [Trp12]-4(6)(Ser) was able to form cation-selective ion channels with two levels of conductance (mainly 250 and occasionally 125 pS) in asolectin planar lipid bilayer, suggesting that appropriate orientation of hydrophobic and hydrophilic residues in amphipathic peptide can simplify channel current levels.

Conformational studies of amphipathic alpha-helical peptides containing an amino acid with a long alkyl chain and their anchoring to lipid bilayer liposomes.

In order to investigate the conformation and orientation of lipid-bound peptides and proteins in the lipid bilayer, basic amphipathic alpha-helical peptides with a long alkyl chain, palmitoyl-(Leu-Ala-Arg-Leu)3-NHCH3 (P-4(3)) and Ac-Leu-Ala-Arg-Leu-Trp-Amy-Arg-Leu-Leu-Ala-Arg-Leu-NHCH3 (Amy-4(3), Amy; alpha-aminomyristic acid) were designed and synthesized. The conformational features and spectroscopic behavior in a buffer solution and in neutral and acidic liposomes were studied by CD, dye-leakage, and fluorescence measurements. The CD data indicated that P-4(3) took an alpha-helical structure in aqueous solution and in neutral and acidic liposomes. On the other hand, Amy-4(3) took a beta-structure in aqueous solution and an alpha-helical structure in neutral and acidic liposomes. The conformational change of Amy-4(3) was confirmed by fluorescence study on lipid titration of the peptide. The dye-leakage experiment showed that both peptides interacted with acidic liposomes to perturb them, but less effectively than Ac-(Leu-Ala-Arg-Leu)3-NHCH3 (4(3)) which has no long alkyl group. Based on these results, a discussion is made concerning the conformation and orientation of peptides in aqueous solution and in the lipid bilayer.

Basic amphipathic helical peptides induce destabilization and fusion of acidic and neutral liposomes.

We have studied the fusion of small unilamellar vesicles composed of egg PC and of a mixture of egg PC plus egg PA using various basic amphipathic peptides. Fusion was monitored by carboxyfluorescein leakage assay, light scattering, membrane intermixing assay, contents mixing assay and electron microscopy. Ac-(L-Leu-L-Ala-L-Arg-L-Leu)3-NHCH3 (peptide 4(3] and Ac-(L-Leu-L-Ala-L-Lys-L-Leu)3-NHCH3 (peptide 4'3), which have high hydrophobic moments, caused transformation of small unilamellar vesicles into larger and relatively homogeneous ones. Ac-(L-Leu-L-Leu-L-Ala-L-Arg-L-Leu)2-NHCH3 (5(2], which has medium hydrophobic moment, induced weak but appreciable fusion, while Ac-(L-Ala-L-Arg-L-Leu)3-NHCH3 (3(3] which has no helical structure did not show any fusion. However, peptides 4(3), 4'3 and 5(2) caused massive leakage of the contents from small unilamellar vesicles. These results indicated that interaction of the peptides with artificial membranes caused extensive perturbation of the lipid bilayer, followed by fusion. The fusogenic capacity of model basic peptides was correlated with the hydrophobic moment of each peptide when the peptides adopted an alpha-helical structure in the presence of acidic liposomes. Peptides 4(3) and 4'3 also showed weak fusogenic ability for neutral liposomes, while 5(2) and 3(3) showed no ability, suggesting that highly amphipathic peptides, such as 4(3), interact weakly but distinctly with neutral liposomes to fuse them.

Solution structure of micelle-bound H5 peptide (427-452): a primary structure corresponding to the pore forming region of the voltage dependent potassium channel.

A 26-mer peptide with the sequence of the pore forming region (residues 427-452) of the Shaker K(+) channel (H5 region) was chemically synthesized. Analyses by CD and two-dimensional 1H NMR spectroscopy were used to investigate the structure of the peptide bound to SDS micelles in solution, which are commonly used in biophysical studies. The tertiary structure of the peptide as a monomer was composed of an alpha-helix (431-438), a turn (439-442), and random coils (427-430, 443-452), and was very similar to that of the pore forming region of the native K(+) channel from Streptomyces lividans determined by X-ray analysis. This result suggests that even an isolated peptide forms a native-like conformation for residues from 431 to 442, depending on its intrinsic amino acid sequence and the surrounding environment.

Orientational behavior of phospholipid membranes with mastoparan studied by 31P solid state NMR.

Solid state 31P NMR spectroscopy was used to study the perturbing effect of the wasp venom peptide mastoparan (MP) on lipid bilayers composed of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylglycerol (DMPG). The 31P chemical shift anisotropy of multilamellar vesicles decreased with increasing peptide concentration, indicating that MP interacts strongly and selectively with the charged DMPG head group. Macroscopically oriented MP-lipid samples between glass plates were studied by 31P NMR as a function of tilt angle. These spectra showed the coexistence of orientation-dependent lamellar signals as well as an isotropic peak, suggesting that MP can induce non-lamellar phases in DMPC/DMPG membranes.