Reduction of membrane fluidity by antibacterial sophoraflavanone G isolated from Sophora exigua.

Sophoraflavanone G (5,7,2',4'-tetrahydroxy-8-lavandulylflavanone) has been referred as a phytochemical with the intensive antibacterial activity. To elucidate the pharmacological mechanism underlying an antibacterial action, sophoraflavanone G was isolated from Sophora exigua, thereafter its effect on membrane fluidity was studied using model membranes and compared with less active naringenin lacking 8-lavandulyl and 2'-hydroxyl groups. Highly purified sophoraflavanone G of 0.05-5 microg/ml, corresponding to the minimum growth inhibitory concentrations against various bacteria, significantly increased fluorescence polarization of the liposomes prepared from 1,2-dipalmitoyl-L-alpha-phosphatidylcholine and 1-palmitoyl-2-oleoyl-L-alpha-phosphatidylcholine. Such increases were found in both liposomes measured with two fluorescent probes to indicate an alteration of membrane fluidity in hydrophilic and hydrophobic regions, suggesting that sophoraflavanone G reduces the fluidity of outer and inner layers of membranes. Although naringenin also showed the membrane effect, it needed concentrations over 2.5 microg/ml to induce a significant reduction of membrane fluidity. Sophoraflavanone G is considered to exert an antibacterial effect by reducing the fluidity of cellular membranes.     

Adenosine 5''-triphosphate-linked transhydrogenase in cytoplasmic membranes of colicin-treated and untreated Escherichia coli.

The adenosine 5'-triphosphate (ATP)-linked transhydrogenase reaction, present in the particulate fractions of Escherichia coli, was previously shown to be inhibited in these fractions when the bacteria were treated with colicins K or El. The purpose of this study was to characterized the ATP-linked transhydrogenase reaction and the colicin-caused inhibition of the reaction in purified cytoplasmic membranes. Particulate fractions from bacteria treated or untreated with colicins were separated on sucrose gradients into cell wall membrane and cytoplasmic membrane fractions. The ATP-linked transhydrogenase reaction was found to be exclusively associated with the cytoplasmic membrane fractions. The reaction was inhibited by carbonylcyanide m-chlorophenlhdrazone, dinitrophenol, N,N'-dicyclohexylcarbodiimide, and trypsin. Although the cytoplasmic membrane fractions were purified from the majoriy of the cell wall membrane and its bound colicins, they showed the inhibitory effects of colicins K and El on the ATP-linked transhydrogenase reaction. The inhibition of ATP-linked transhydrogenase reaction induced by the colicin could not be reversed by subjection the isolated membranes to a variety of physical and chemical treatments. Cytoplasmic membranes depleted of energy-transducing adenosine triphosphatase ATPase) complex (coupling factor) lost the ATP-linked transhydrogenase activity. The ATPase complexes isolated from membranes of bacteria treated or untreated with colicins El or K reconstituted high levels of ATP-linded transhydrogenase activity to depleted membranes of untreated bacteria. The same ATPase complexes reconstituted low levels of activity to depleted membranes of the treated bacteria.     

Effect of ionic and colloid gold on ATP-hydrolase fermentative systems in the membrane of Bacillus sp. B4253 and Bacillus sp. B4851

Accumulation of gold in cells of Bacillus sp. B4253 can be directly or indirectly connected with activity of bacteria plasma membrane basal Mg2+-ATPase. Therefore this work deals with a comparative analysis of kinetic properties of plasma membrane basal azide-resistant Mg2+-dependent ATP-hydrolase activity of B. sp. B4253 and B. sp. B4851 capable to gold accumulation and not capable to this process, accordingly. It is shown, that by a number of kinetic parameters - specific fermentative activity, initial speed of reaction of hydrolysis ATP (V0), Mixaelis constant (Km), the maximal initial speed by Mg2+ (V(Mg)) and by ATP (V(ATP)), optimum concentration of ATP ([ATP]opt), pHmax, sensitivity to action of the thapsigargine and eosine Y - bacteria membranes basal Mg2+-ATPase activity accumulating gold, and the bacteria not capable to this process, are identical. But by some parameters they differ: Mg2+-ATPase activity of membranes of the bacteria which do not accumulate gold, has three times greater affinity for Mg ions and smaller value [Mg]opt. The inhibition effect of ionic gold (10(-4)-3x10(-4) M) is shown on azide-sensitive (H+-ATPase) and azide-resistant (Mg2+-ATPase) components Mg2+-dependent ATP-hydrolase activity in fraction of plasma membranes of microorganisms Bacillus accumulating gold, and not capable to this process. Colloid gold (0.0002-4 microg/ml) stimulates activity of H+-ATPase and Mg2+-ATPase in a membrane of the bacteria accumulating gold 1.5-2 times, and does not influence activity of ATPases of a membrane of the bacteria which do. not accumulate gold.     

NH2-terminal signals in ATP7B Cu-ATPase mediate its Cu-dependent anterograde traffic in polarized hepatic cells

Cu is an essential cofactor of cellular proteins but is toxic in its free state. The hepatic Cu-ATPase ATP7B has two functions in Cu homeostasis: it loads Cu+ onto newly synthesized apoceruloplasmin in the secretory pathway, thereby activating the plasma protein; and it participates in the excretion of excess Cu+ into the bile. To carry out these two functions, the membrane protein responds to changes in intracellular Cu levels by cycling between the Golgi and apical region. We used polarized hepatic WIF-B cells and high-resolution confocal microscopy to map the itinerary of endogenous and exogenous ATP7B under different Cu conditions. In Cu-depleted cells, ATP7B resided in a post-trans-Golgi network compartment that also contained syntaxin 6, whereas in Cu-loaded cells, the protein relocated to unique vesicles very near to the apical plasma membrane as well as the membrane itself. To determine the role of ATP7B's cytoplasmic NH2 terminus in regulating its intracellular movements, we generated seven mutations/deletions in this large [approximately 650 amino acid (AA)] domain and analyzed the Cu-dependent behavior of the mutant ATP7B proteins in WIF-B cells. Truncation of the ATP7B NH2 terminus up to the fifth copper-binding domain (CBD5) yielded an active ATPase that was insensitive to cellular Cu levels and constitutively trafficked to the opposite (basolateral) plasma membrane domain. Fusion of the NH2-terminal 63 AA of ATP7B to the truncated protein restored both its Cu responsiveness and correct intracellular targeting. These results indicate that important targeting information is contained in this relatively short sequence, which is absent from the related CuATPase, ATP7A.     

Deciphering the role of Burkholderia cenocepacia membrane proteins in antimicrobial properties of chitosan

Chitosan, a versatile derivative of chitin, is widely used as an antimicrobial agent either alone or mixed with other natural polymers. Burkholderia cenocepacia is a multidrug-resistant bacteria and difficult to eradicate. Our previous studies shown that chitosan had strong antibacterial activity against B. cenocepacia. In the current study, we have investigated the molecular aspects for the susceptibility of B. cenocepacia in response to chitosan antibacterial activity. We have conducted RNA expression analysis of drug efflux system by RT-PCR, membrane protein profiling by SDS–PAGE, and by LC-MS/MS analysis following the validation of selected membrane proteins by real-time PCR analysis. By RT-PCR analysis, it was found that orf3, orf9, and orf13 were expressed at detectable levels, which were similar to control, while rest of the orf did not express. Moreover, shotgun proteomics analysis revealed 21 proteins in chitosan-treated cells and 16 proteins in control. Among them 4 proteins were detected as shared proteins under control and chitosan-treated cells and 17 proteins as uniquely identified proteins under chitosan-treated cells. Among the catalog of uniquely identified proteins, there were proteins involved in electron transport chain and ATP synthase, metabolism of carbohydrates and adaptation to atypical conditions proteins which indicate that utilization and pattern of chitosan is diverse which might be responsible for its antibacterial effects on bacteria. Moreover, our results showed that RND drug efflux system, which display the ability to transport a variety of structurally unrelated drugs from a cell and consequently are capable of conferring resistance to a diverse range of chemotherapeutic agents, was not determined to play its role in response to chitosan. It might be lipopolysaccharides interaction with chitosan resulted in the destabilization of membrane protein to membrane lyses to cell death. Membrane proteome analysis were also validated by RT-qPCR analysis, which corroborated our results that of membrane proteins.     

Naphthalene association and uptake in Pseudomonas putida.

Two methods for bacterial membrane transport, filtration and flow dialysis, were used to study cellular association of Pseudomonas putida with naphthalene. It is not technically possible to determine the exact cellular or vesicular location of the naphthalene, and because it is hydrophobic, it could be at the membrane(s) rather than inside the cells. As an index of naphthalene having crossed the inner membrane we used the intracellular formation of its first catabolite. An energized membrane or ATP was not essential for association or movement into the cell. Evidence for a nonspecific association and a movement into cells by simple diffusion are the lack of saturation of association, an absence of inhibition of association by protein inhibitors and structural analogs, and the passage of naphthalene through cell membranes in the presence of iodoacetamide. Specific naphthalene metabolism gene expression was not required for association.     

In vitro antibacterial activity and antifungal mode of action of flocculosin, a membrane-active cellobiose lipid

Aims:  To investigate the in vitro antibacterial activity and antifungal mode of action of flocculosin, a cellobiose lipid produced by Pseudozyma flocculosa .
Methods and Results:  When tested against clinical bacterial isolates, the compound was particularly active against Gram-positive bacteria and its effect was not mitigated against isolates known as resistant to other antibiotics. The antifungal activity of flocculosin was found to be rapid and concentration-dependent. At lethal concentrations against Candida albicans , flocculosin caused a rapid leakage of intracellular potassium and inhibited acidification of the medium by plasma membrane ATPases suggesting a physical rather than a biochemical effect. TEM observations of cells exposed 6 h to flocculosin revealed disrupted membranes and disorganized mitochondria.
Conclusions:  Data obtained in this study confirm that flocculosin acts by disrupting the membrane surface of sensitive micro-organisms.
Significance and Impact of the Study:  The elucidation of an antifungal mode of action of flocculosin can be exploited in furthering its antimicrobial potential against fungi and bacteria whose cell membranes are particularly sensitive to the action of the molecule.     

Dual mechanism of bacterial lethality for a cationic sequence-random copolymer that mimics host-defense antimicrobial peptides

Flexible sequence-random polymers containing cationic and lipophilic subunits that act as functional mimics of host-defense peptides have recently been reported. We used bacteria and lipid vesicles to study one such polymer, having an average length of 21 residues, that is active against both Gram-positive and Gram-negative bacteria. At low concentrations, this polymer is able to permeabilize model anionic membranes that mimic the lipid composition of Escherichia coli, Staphylococcus aureus, or Bacillus subtilis but is ineffective against model zwitterionic membranes, which explains its low hemolytic activity. The polymer is capable of binding to negatively charged vesicles, inducing segregation of anionic lipids. The appearance of anionic lipid-rich domains results in formation of phase-boundary defects through which leakage can occur. We had earlier proposed such a mechanism of membrane disruption for another antimicrobial agent. Experiments with the mutant E. coli ML-35p indicate that permeabilization is biphasic: at low concentrations, the polymer permeabilizes the outer and inner membranes; at higher polymer concentrations, permeabilization of the outer membrane is progressively diminished, while the inner membrane remains unaffected. Experiments with wild-type E. coli K12 show that the polymer blocks passage of solutes into the intermembrane space at high concentrations. Cell membrane integrity in E. coli K12 and S. aureus exhibits biphasic dependence on polymer concentration. Isothermal titration calorimetry indicates that the polymer associates with the negatively charged lipopolysaccharide of Gram-negative bacteria and with the lipoteichoic acid of Gram-positive bacteria. We propose that this polymer has two mechanisms of antibacterial action, one predominating at low concentrations of polymer and the other predominating at high concentrations.     

Adenosine triphosphatase activity of mycoplasma membranes

Rottem, Shlomo (Hebrew University, Jerusalem, Israel), and Shmuel Razin. Adenosine triphosphatase activity of mycoplasma membranes. J. Bacteriol. 92:714-722. 1966.-Adenosine triphosphatase activity of Mycoplasma laidlawii, M. gallisepticum, and Mycoplasma sp. strain 14 was confined to the cell membrane. The enzymatic activity was dependent on magnesium, but was not activated by sodium and potassium. Ouabain did not inhibit the adenosine triphosphatase activity of the mycoplasmas, and did not interfere with the active accumulation of potassium by M. laidlawii cells. Sulfhydryl-blocking reagents and fluoride inhibited the enzymatic activity, whereas 2,4-dinitrophenol was without any effect. Membranes of M. laidlawii hydrolyzed other nucleotide triphosphates and adenosine diphosphate (ADP), but at a lower rate than adenosine triphosphate (ATP). Nucleoside-2'-(3')-phosphates, ribose-5-phosphate, glucose-6-phosphate, and pyrophosphate were not hydrolyzed by the membrane preparations. It seems that the enzyme(s) involved in ATP hydrolysis by M. laidlawii membranes is strongly bound to the membrane subunits, which would account for the failure to purify the enzyme by protein fractionation techniques. The adenosine triphosphatase activity of mycoplasma membranes resembles in its properties that of similar enzymes studied in bacteria. The mycoplasma enzyme(s) seems to differ from the adenosine triphosphatase associated with ion transport in mammalian cell membranes and from mitochondrial adenosine triphosphatase.     

Antimicrobial properties of analgesic kyotorphin peptides unraveled through atomic force microscopy

Antimicrobial peptides (AMPs) are promising candidates as alternatives to conventional antibiotics for the treatment of resistant pathogens. In the last decades, new AMPs have been found from the cleavage of intact proteins with no antibacterial activity themselves. Bovine hemoglobin hydrolysis, for instance, results in AMPs and the minimal antimicrobial peptide sequence was defined as Tyr-Arg plus a positively charged amino acid residue. The Tyr-Arg dipeptide alone, known as kyotorphin (KTP), is an endogenous analgesic neuropeptide but has no antimicrobial activity itself. In previous studies new KTP derivatives combining C-terminal amidation and Ibuprofen (Ib) - KTP-NH(2), IbKTP, IbKTP-NH(2) - were designed in order to improve KTP brain targeting. Those modifications succeeded in enhancing peptide-cell membrane affinity towards fluid anionic lipids and higher analgesic activity after systemic injection resulted therefrom. Here, we investigated if this affinity for anionic lipid membranes also translates into antimicrobial activity because bacteria have anionic membranes. Atomic force microscopy revealed that KTP derivatives perturbed Staphylococcus aureus membrane structure by inducing membrane blebbing, disruption and lysis. In addition, these peptides bind to red blood cells but are non-hemolytic. From the KTP derivatives tested, amidated KTP proves to be the most active antibacterial agent. The combination of analgesia and antibacterial activities with absence of toxicity is highly appealing from the clinical point of view and broadens the therapeutic potential and application of kyotorphin peptides.     

The antibacterial activity of phospholipase A2 type IIA is regulated by the cooperative lipid chain melting behavior in Staphylococcus aureus

As one of its primary physiological functions, sPLA₂-IIA appears to act as an antibacterial agent. In particular, sPLA₂-IIA shows high activity towards Gram-positive bacteria such as Staphylococcus aureus (S. aureus). This antibacterial activity results from the preference of the enzyme towards membranes enriched in anionic lipids, which is a common feature of bacterial membranes. An intriguing aspect observed in a variety of bacterial membranes is the presence of a broad but cooperative lipid chain melting event where the lipids in the membrane transition from a solid-ordered (so) into a liquid-disordered (ld) state close to physiological temperatures. It is known that the enzyme is sensitive to the level of lipid packing, which changes sharply between the so and the ld states. Therefore, it would be expected that the enzyme activity is regulated by the bacterial membrane thermotropic behavior. We determine by FTIR the thermotropic lipid chain melting behavior of S. aureus and find that the activity of sPLA₂-IIA drops sharply in the so state. The activity of the enzyme is also evaluated in terms of its effects on cell viability, showing that cell survival increases when the bacterial membrane is in the so state during enzyme exposure. These results point to a mechanism by which bacteria can develop increased resistance towards antibacterial agents that act on the membrane through a cooperative increase in the order of the lipid chains. These results show that the physical behavior of the bacterial membrane can play an important role in regulating physiological function in an in vivo system.     

Redox modulation of the response of NADH oxidase activity of rat liver plasma membranes to cyclic AMP plus ATP

NADH oxidase activity of rat liver plasma membranes was inhibited by lowconcentrations (1-100 nM) of ATP. The inhibition was amplified by additionof nanomolar concentrations (0.1-10) of cyclic AMP. The inhibition wascomplex and related to a marked increase in the Km for NADH at high NADHconcentrations together with a concomitant decrease in the Vmax. In theabsence of added or residual ATP, cyclic AMP was without effect. Theresponse of cyclic AMP + ATP was inhibited by low concentrations of theselective inhibitor of cyclic AMP-dependent protein kinase, H-89 but not bystaurosporin. The Vmax but not the Km was modified by treating the plasmamembranes with a mild oxidizing agent, N-chlorosuccinamide, or with thereducing agent, dithiothreitol. In the presence of dithiothreitol, the Vmaxwas reduced by cyclic AMP + ATP. In contrast, in the presence ofN-chlorosuccinamide, the Vmax was increased by cyclic AMP + ATP relative tocyclic AMP + ATP alone. Thus, the effect of cyclic AMP + ATP on the Vmaxcould be either an increase or a decrease depending on whether the membraneswere oxidized or reduced. The results demonstrate regulation of NADH oxidaseactivity of rat liver plasma membranes through cyclic AMP-mediatedphosphorylation by membrane-located protein kinase activities where thefinal response is dependent on the oxidation-reduction status of the plasmamembranes.     

Constitutive ERK MAPK activity regulates macrophage ATP production and mitochondrial integrity

A unique feature of human alveolar macrophages is their prolonged survival in the face of a stressful environment. We have shown previously that the ERK MAPK is constitutively active in these cells and is important in prolonging cell survival. This study examines the role of the ERK pathway in maintaining mitochondrial energy production. The data demonstrate that ATP levels in alveolar macrophages depend on intact mitochondria and optimal functioning of the electron transport chain. Significant levels of MEK and ERK localize to the mitochondria and inhibition of ERK activity induces an early and profound depletion in cellular ATP coincident with a loss of mitochondrial transmembrane potential. The effect of ERK suppression on ATP levels was specific, since it did not occur with PI3K/Akt, p38, or JNK suppression. ERK inhibition led to cytosolic release of mitochondrial proteins and caspase activation. Both ERK inhibition and mitochondrial blockers induced loss of plasma membrane permeability and cell death. The cell death induced by ERK inhibition had hallmarks of both apoptotic (caspase activation) and necrotic (ATP loss) cell death. By blocking ERK inhibition-induced reactive oxygen species, caspase activation was prevented, although necrotic pathways continued to induce cell death. This suggests that mitochondrial dysfunction caused by ERK inhibition generates both apoptotic and necrotic cell death-inducing pathways. As a composite, these data demonstrate a novel mitochondrial role for ERK in maintaining mitochondrial membrane potential and ATP production in human alveolar macrophages.     

Identification of antibacterial mechanism of L-amino acid oxidase derived from Trichoderma harzianum ETS 323

Although L-amino oxidase (LAAO; EC 1.4.3.2) has been reported to be a potent antibacterial agent, the mechanism responsible for its antibacterial activity has not been identified. The present study aimed to identify the mechanism responsible for the antibacterial activity of Th-LAAO, an LAAO recently isolated from the extracellular proteins of Trichoderma harzianum ETS 323, at the same time as elucidating the nature of this enzyme. The results obtained indicate that the enzyme activity and structure of Th-LAAO are stable at pH 6-8 and less stable at both pH 4-5.5 and pH 9. At pH 7.0, the optimum temperature for Th-LAAO was found to be 40 °C, comprising the temperature at which enzymatic activity is greatest, with enzymatic activity deceasing with further increases in temperature as a result of thermal denaturation of the enzyme, leading to partial denaturation at 50 °C. The results obtained by confocal microscopy and flow cytometry indicate that Th-LAAO interacts with bacteria to cause membrane permeabilization, and this interaction may be promoted by the amphipathic sequence in Th-LAAO and other cytotoxic LAAOs located at the N-terminus. The findings of increased exogenous H(2) O(2) production and reactive oxidative species accumulation in Th-LAAO-treated bacteria indicate that reactive oxidative species accumulation may trigger forms of cell damage, including lipid peroxidation and DNA strand breakage that results in bacterial growth inhibition. Taken together, the results indicate that the processes of bacterial interaction, membrane permeabilization and H(2)O(2) production are involved in the mechanism responsible for the antibacterial activity of Th-LAAO.     

3-D structural and functional characterization of the purified KATP channel complex Kir6.2-SUR1

ATP-sensitive potassium (K(ATP)) channels conduct potassium ions across cell membranes and thereby couple cellular energy metabolism to membrane electrical activity. Here, we report the heterologous expression and purification of a functionally active K(ATP) channel complex composed of pore-forming Kir6.2 and regulatory SUR1 subunits, and determination of its structure at 18 A resolution by single-particle electron microscopy. The purified channel shows ATP-ase activity similar to that of ATP-binding cassette proteins related to SUR1, and supports Rb(+) fluxes when reconstituted into liposomes. It has a compact structure, with four SUR1 subunits embracing a central Kir6.2 tetramer in both transmembrane and cytosolic domains. A cleft between adjacent SUR1s provides a route by which ATP may access its binding site on Kir6.2. The nucleotide-binding domains of adjacent SUR1 appear to interact, and form a large docking platform for cytosolic proteins. The structure, in combination with molecular modelling, suggests how SUR1 interacts with Kir6.2.     

MacB ABC transporter is a dimer whose ATPase activity and macrolide-binding capacity are regulated by the membrane fusion protein MacA

Gram-negative bacteria utilize specialized machinery to translocate drugs and protein toxins across the inner and outer membranes, consisting of a tripartite complex composed of an inner membrane secondary or primary active transporter (IMP), a periplasmic membrane fusion protein, and an outer membrane channel. We have investigated the assembly and function of the MacAB/TolC system that confers resistance to macrolides in Escherichia coli. The membrane fusion protein MacA not only stabilizes the tripartite assembly by interacting with both the inner membrane protein MacB and the outer membrane protein TolC, but also has a role in regulating the function of MacB, apparently increasing its affinity for both erythromycin and ATP. Analysis of the kinetic behavior of ATP hydrolysis indicated that MacA promotes and stabilizes the ATP-binding form of the MacB transporter. For the first time, we have established unambiguously the dimeric nature of a noncanonic ABC transporter, MacB that has an N-terminal nucleotide binding domain, by means of nondissociating mass spectrometry, analytical ultracentrifugation, and atomic force microscopy. Structural studies of ABC transporters indicate that ATP is bound between a pair of nucleotide binding domains to stabilize a conformation in which the substrate-binding site is outward-facing. Consequently, our data suggest that in the presence of ATP the same conformation of MacB is promoted and stabilized by MacA. Thus, MacA would facilitate the delivery of drugs by MacB to TolC by enhancing the binding of drugs to it and inducing a conformation of MacB that is primed and competent for binding TolC. Our structural studies are an important first step in understanding how the tripartite complex is assembled.     

Application of zinc oxide and sodium alginate for biofouling mitigation in a membrane bioreactor treating urban wastewater

Abstract

This research aimed to mitigate fouling in membrane bioreactors (MBR) through concurrent usage of zinc oxide as an antibacterial agent (A) and sodium alginate as a hydrophilic agent (H) within a polyacrylonitrile membrane (PM) structure. The antibacterial polymeric membranes (APM) and antibacterial hydrophilic polymeric membranes (AHPM) synthesized showed a higher porosity, mechanical strength and bacterial inhibition zone, and a lower contact angle in comparison with PM membranes. EDS, SEM and AFM analyses were used to characterize the chemical, structural, and morphological properties of PM, APM, and AHPM. The flux of PM, APM, and AHPM in MBR was 37, 48, and 51?l m^?2 h^?1 and COD removal was 81, 93.5, and 96.7%, respectively. After MBR operation for 35?days in an urban wastewater treatment, only 50% of the flux of PM was recovered, while the antibacterial and hydrophilic agents yielded a flux recovery of 72.7 and 100% for APM and AHPM, respectively.     

Novel starch/chitosan blending membrane: Antibacterial,permeable and mechanical properties

Antibacterial membranes were prepared from a mixture of hydrolyzed starch and chitosan. Glycerin was incorporated in the membranes to as plasticizer agent. The effects of component ratio on the mechanical and permeable properties of the prepared membranes were investigated. The elongation-at-break and water vapor transmission rate of starch/chitosan blending membranes were largely improved compared with each single component due to the interaction formed between the hydroxyl groups of starch and the amino ones of chitosan, which was confirmed by FT-IR characterizations. With the help of optical microscope, the influence of component ratio on the morphologies of starch/chitosan membranes was systematically investigated. It comes to a conclusion that extreme low or high starch content will cause an asymmetric membrane surface. To prove the antibacterial activity of obtained membranes, Escherichia coli (E. coli) was chosen as the target bacteria via optical density method. The resulted starch/chitosan membranes exhibited an outstanding antibacterial activity against E. coli.     

Antibacterial activity of peptides derived from the C-terminal region of a hemolytic lectin, CEL-III, from the marine invertebrate Cucumaria echinata

Several synthetic peptides derived from the C-terminal domain sequence of a hemolytic lectin, CEL-III, were examined as to their action on bacteria and artificial lipid membranes. Peptide P332 (KGVIFAKASVSVKVTASLSK-NH(2)), corresponding to the sequence from residue 332, exhibited strong antibacterial activity toward Gram-positive bacteria. Replacement of each Lys in P332 by Ala markedly decreased the activity. However, when all Lys were replaced by Arg, the antibacterial activity increased, indicating the importance of positively charged residues at these positions. Replacement of Val by Leu also led to higher antibacterial activity, especially toward Gram-negative bacteria. The antibacterial activity of these peptides was correlated with their membrane-permeabilizing activity toward the bacterial inner membrane and artificial lipid vesicles, indicating that the antibacterial action is due to perturbation of bacterial cell membranes, leading to enhancement of their permeability. These results also suggest that the hydrophobic region of CEL-III, from which P332 and its analogs were derived, may play some role in the interaction with target cell membranes to trigger hemolysis.