<i>Insilico</i> Anticancer Peptide Prediction from <i>Curcuma longa</i>

Cancer is the second biggest cause of death globally, and the use of therapeutic peptides to specifically target and destroy cancer cells has gotten much interest. Cancer peptides or vaccinations are utilized to treat cancer nowadays, apart from chemotherapy, which has significant discomfort, side effects and costly. It is time demanding to identify and predict potential anticancer peptides using computational biology approaches. Thus, 3-D molecular modeling is being used to find possible ACP candidates. In this research, Curcuma longa has predicted peptide sequences were docked on breast cancer receptors and used a molecular docking technique to assess the anticipated peptides’ binding affinities to MHC molecules. A similar approach was utilized to simulate the interactions of the chosen peptide with the TCR. Additionally, the Pep10 LIRQHVASNIGIAKSKIREPIV was examined, and our findings indicated interaction with MHC classes I and II. However, the maximum binding energy was obtained with TCR at 695.61, giving strength through eight hydrogen bonds. Similarly, the Pep20, GAIIGNRKIKLQPHIIIRID, the projected, has the most significant overall binding energy with MHC classes I and II but a lower global E total value with TCR, namely −600.97 kj/Mol, and also four hydrogen bonds. This research could lead to the development of novel anticancer drugs based on the anticancer activity of the Curcuma longa medicinal plant.     

PEP and CADY-mediated delivery of fluorescent peptides and proteins into living cells

Cell-penetrating peptides (CPPs) constitute a family of peptides with the characteristic ability to cross biological membranes and deliver cargo into the intracellular milieu. Several CPPs have been proposed for delivery of polypeptides and proteins into cells through either of two strategies: covalent or complexed in a non-covalent fashion. Members of the PEP family are primary amphipathic peptides which have been shown to deliver peptides and proteins into a wide variety of cells through formation of non-covalent complexes. CADY is a secondary amphipathic peptide which has been demonstrated to deliver short nucleic acids, in particular siRNA with high efficiency. Here we review the characteristics of the PEP and CADY carriers and describe a novel derivative of CADY termed CADY2, which also presents sequence similarities to Pep1. We have compared Pep1, CADY and CADY2 in their efficiency to interact with and internalize short fluorogenic peptides and proteins into cultured cells, and provide evidence that CADY2 can interact with proteins and peptides and deliver them efficiently into living cells, similar to Pep1, but in contrast to CADY which is unable to deliver any peptide, even short negatively charged peptides. This is the first study to investigate the influence of the cargo on the interactions between PEP and CADY carriers, thereby providing novel insights into the physicochemical parameters underlying interactions and cellular uptake of peptides and proteins by these non-covalent CPPs.     

Identification and characterization of a multispecific monoclonal antibody G2 against chicken prion protein

We previously generated a monoclonal antibody (mAb), G2, by immunizing mice with Residues 174–247 of the chicken prion protein (ChPrP^C). In this study, we found that G2 possessed an extremely unusual characteristic for a mAb; in particular, it could react with at least three proteins other than ChPrP^C, the original antigenic protein. We immunoscreened a complementary DNA library from chicken brain DNA and found three proteins (SEPT3, ATP6V1C1, and C6H10orf76) that reacts with G2. There were no regions of amino acid sequence similarity between ChPrP^C and SEPT3, ATP6V1C1, or C6H10orf76. We selected ATP6V1C1 as a representative of the three proteins and identified the epitope within ATP6V1C1 that reacts with G2. The amino acid sequence of the G2 epitope within ATP6V1C1 (Pep8) was not related to the G2 epitope within ChPrP^C (Pep18mer). However, enzyme-linked immunosorbent assay, surface plasmon resonance (SPR), and isothermal titration calorimetry (ITC) experiments indicated that these two peptides have similar binding affinity for G2. The apparent K_D values of Pep18mer and Pep8 obtained from SPR experiments were 2.9 × 10⁻⁸ and 1.6 × 10⁻⁸ M, respectively. Antibody inhibition test using each peptide indicated that the binding sites of the two different peptides overlapped each other. We observed that these two peptides substantially differed in several binding characteristics. Based on the SPR experiments, the association and dissociation rate constants of Pep18mer were higher than those of Pep8. A clear difference was also observed in ITC experiments. These differences may be explained by G2 adopting different binding conformations and undergoing different binding pathways.     

A lactoferrin-derived peptide with cationic residues concentrated in a region of its helical structure induces necrotic cell death in a leukemic cell line (HL-60).

Model studies have shown that peptides derived from the N-terminal region of bovine lactoferrin (Lf-B) exhibit antitumor activity against certain cell lines. This activity is due primarily to the peptides' apoptogenic effect. Several reports indicate that cationic residues clustered in two regions of the peptide sequence can be shuffled into one region and thereby increase cytotoxic activity, although the mechanism of this enhanced cytotoxic effect has not been clarified. In this paper, we considered several parameters that determine the mode of cell death after exposure to a native Lf-B derived peptide (Pep1, residues 17-34), and a modified peptide (mPep1) wherein the cationic residues of Pep1 are clustered in a single region of its helical structure. We found that the cytotoxic activity of mPep1 was about 9.6 fold-higher than that of Pep1 against HL-60 cells, as determined by the 3-(4,5-dimethylthiazol-2-yl)-5(3-carboxymethonyphenol)-2-(4-sulfophenyl)-2 H-tetrazolium (MTS) assay. In investigating the expression of phosphatidylserine, we observed that the native peptide (Pep1) caused both apoptotic cell death and necrotic cell death, depending on the concentration of the peptide. In contrast, the action of mPep1 was exclusively characteristic of necrotic cell death. This observation was further confirmed by agarose gel electrophoresis, in which clear ladder-like DNA bands were observed from cells exposed to Pep1, whereas DNA from cells treated with mPep1 produced a smeared pattern. We extended the study by investigating the release of mitochondrial cytochrome c into the cytosol, and the activation of caspase-3; both peptides caused the release of cytochrome c into the cytosol, and the activation of caspase-3.These results suggest that Pep1 may kill cancer cells by activating an apoptosis-inducing pathway, whereas mPep1 causes necrotic cell death by destroying cellular membrane structure notwithstanding sharing some cellular events with apoptotic cell death. Copyright (c) 2008 European Peptide Society and John Wiley & Sons, Ltd.     

Synthesis,characterization, and biological activity of poly(arginine)‐derived cancer‐targeting peptides in HepG2 liver cancer cells

The solid‐phase synthesis, structural characterization, and biological evaluation of a small library of cancer‐targeting peptides have been determined in HepG2 hepatoblastoma cells. These peptides are based on the highly specific Pep42 motif, which has been shown to target the glucose‐regulated protein 78 receptors overexpressed and exclusively localized on the cell surface of tumors. In this study, Pep42 was designed to contain varying lengths (3–12) of poly(arginine) sequences to assess their influence on peptide structure and biology. Peptides were effectively synthesized by 9‐fluorenylmethoxycarbonyl‐based solid‐phase peptide synthesis, in which the use of a poly(ethylene glycol) resin provided good yields (14–46%) and crude purities >95% as analyzed by liquid chromatography–mass spectrometry. Peptide structure and biophysical properties were investigated using circular dichroism spectroscopy. Interestingly, peptides displayed secondary structures that were contingent on solvent and length of the poly(arginine) sequences. Peptides exhibited helical and turn conformations, while retaining significant thermal stability. Structure–activity relationship studies conducted by flow cytometry and confocal microscopy revealed that the poly(arginine) derived Pep42 sequences maintained glucose‐regulated protein 78 binding on HepG2 cells while exhibiting cell translocation activity that was contingent on the length of the poly(arginine) strand. In single dose (0.15 mM) and dose‐response (0–1.5 mM) cell viability assays, peptides were found to be nontoxic in human HepG2 liver cancer cells, illustrating their potential as safe cancer‐targeting delivery agents. Copyright © 2014 European Peptide Society and John Wiley & Sons, Ltd.     

Protease-activated receptor-1 in human lung fibroblasts mediates a negative feedback downregulation via prostaglandin E2

Among the four protease-activated receptors (PARs), PAR-1 plays an important role in normal lung functioning and in the development of lung diseases, including fibrosis. We compared the expression and functional activity of PARs in normal and fibrotic human lung fibroblasts. Both normal and fibrotic cells express PAR-1, -2, and -3, with PAR-2 showing the lowest level. There was no significant difference between normal and fibrotic fibroblasts in expression levels of PAR-1 and PAR-3, whereas a fourfold higher expression level of PAR-2 was observed in fibrotic cells compared with normal cells. Ca(2+) imaging studies revealed apparently only PAR-1-induced Ca(2+) signaling in lung fibroblasts. PAR-1 agonists, thrombin and synthetic activating peptide, induced concentration-dependent Ca(2+) mobilization with EC(50) values of 5 nM and 1 microM, respectively. The neutrophil protease cathepsin G produced a transient Ca(2+) response followed by disabling PAR-1, whereas elastase did not affect Ca(2+) level. PAR-1 activation by thrombin or receptor-activating peptide downregulated expression of all three PARs in lung fibroblasts, with maximal effect at 3-6 h, whereas expression returned toward basal level after 24 h. Furthermore, PAR-1 agonists dose dependently increased PGE(2) secretion from lung fibroblasts and induction of cyclooxygenase-2 expression. We then found that PGE(2) downregulated expression of all three PARs. The effect of PGE(2) was continuously growing with time. Furthermore, PGE(2) exerts its effect through the EP2 receptor that was confirmed using the selective EP2 agonist butaprost. This novel autocrine feedback mechanism of PGE(2) in lung fibroblasts seems to be an important regulator in lung physiology and pathology.     

Crystal structures of MAP kinase p38 complexed to the docking sites on its nuclear substrate MEF2A and activator MKK3b

The structures of the MAP kinase p38 in complex with docking site peptides containing a phi(A)-X-phi(B) motif, derived from substrate MEF2A and activating enzyme MKK3b, have been solved. The peptides bind to the same site in the C-terminal domain of the kinase, which is both outside the active site and distinct from the "CD" domain previously implicated in docking site interactions. Mutational analysis on the interaction of p38 with the docking sites supports the crystallographic models and has uncovered two novel residues on the docking groove that are critical for binding. The two peptides induce similar large conformational changes local to the peptide binding groove. The peptides also induce unexpected and different conformational changes in the active site, as well as structural disorder in the phosphorylation lip.     

Inhibition of plant-pathogenic bacteria by short synthetic cecropin A-melittin hybrid peptides

Short peptides of 11 residues were synthesized and tested against the economically important plant pathogenic bacteria Erwinia amylovora, Pseudomonas syringae, and Xanthomonas vesicatoria and compared to the previously described peptide Pep3 (WKLFKKILKVL-NH(2)). The antimicrobial activity of Pep3 and 22 analogues was evaluated in terms of the MIC and the 50% effective dose (ED(50)) for growth. Peptide cytotoxicity against human red blood cells and peptide stability toward protease degradation were also determined. Pep3 and several analogues inhibited growth of the three pathogens and had a bactericidal effect at low micromolar concentrations (ED(50) of 1.3 to 7.3 microM). One of the analogues consisting of a replacement of both Trp and Val with Lys and Phe, respectively, resulted in a peptide with improved bactericidal activity and minimized cytotoxicity and susceptibility to protease degradation compared to Pep3. The best analogues can be considered as potential lead compounds for the development of new antimicrobial agents for use in plant protection either as components of pesticides or expressed in transgenic plants.     

Selection and target-site mapping of peptides inhibiting HCV NS5B polymerase using phage display

A series of peptides binding to the HCV NS5B polymerase was selected from phage display peptide libraries. A conserved motif of Ser-Arg-X-Arg/Leu was identified among the selected peptides, and Pep2 (Trp-Ser-Arg-Pro-Arg- Ser-Leu) was chosen for further characterization. The binding of Pep2 to HCV NS5B in vivo was shown by a yeast twohybrid assay and by subcellular colocalization analysis using immunofluorescence confocal microscopy. The in vitro interaction was also confirmed by GST pulldown assay. The replication of the HCV 1b subgenomic replicon was efficiently inhibited by the presence of the peptide. By using a subtractive biopanning against Pep2, the binding site of the peptide was mapped at the pocket of Pro388 to Pro391 in the thumb subdomain of the polymerase. A yeast two-hybrid analysis using Pro388Ala and Pro391Ala mutants of NS5B confirmed the binding.     

Identification of peptides that antagonize formyl peptide receptor-like 1-mediated signaling

Formyl peptide receptor-like 1 (FPRL1) is an important classical chemoattractant receptor that is expressed in phagocytic cells in the peripheral blood and brain. Recently, various novel agonists have been identified from several origins, such as host-derived molecules. Activation of FPRL1 is closely related to inflammatory responses in the host defense mechanism and neurodegenerative disorders. In the present study we identified several novel peptides by screening hexapeptide libraries that inhibit the binding of one of FPRL1's agonists (Trp-Lys-Tyr-Met-Val-D-Met-CONH(2) (WKYMVm)) to its specific receptor, FPRL1, in RBL-2H3 cells. Among the novel peptides, Trp-Arg-Trp-Trp-Trp-Trp-CONH(2) (WRWWWW (WRW(4))) showed the most potent activity in terms of inhibiting WKYMVm binding to FPRL1. We also found that WRW(4) inhibited the activation of FPRL1 by WKYMVm, resulting in the complete inhibition of the intracellular calcium increase, extracellular signal-regulated kinase activation, and chemotactic migration of cells toward WKYMVm. For the receptor specificity of WRW(4) to the FPR family, we observed that WRW(4) specifically inhibit the increase in intracellular calcium by the FPRL1 agonists MMK-1, amyloid beta42 (Abeta42) peptide, and F peptide, but not by the FPR agonist, fMLF. To investigate the effect of WRW(4) on endogenous FPRL1 ligand-induced cellular responses, we examined its effect on Abeta42 peptide in human neutrophils. Abeta42 peptide-induced superoxide generation and chemotactic migration of neutrophils were inhibited by WRW(4), which also completely inhibited the internalization of Abeta42 peptide in human macrophages. WRW(4) is the first specific FPRL1 antagonist and is expected to be useful in the study of FPRL1 signaling and in the development of drugs against FPRL1-related diseases.     

Integrin-dependent interaction of human vascular endothelial cells on biomimetic peptide surfactant polymers

Biomimetic surfactant polymers designed by molecular grafting of pendant RGD peptides (Pep) and dextran oligosaccharides (Dex) in different ratios onto the backbone of poly(vinyl amine) (PVAm) were examined for their ability to promote endothelial cell (EC) growth. Adhesion, formation of focal contacts, and expression of integrin receptors were examined in EC seeded onto a series of novel surfactants containing 100% dextran (PVAm[Pep (0%)]) to 100% peptide (PVAm[Pep (100%)]) compared to fibronectin control. Interaction of EC on polymer was specific, as soluble GRGDSP, but not GRGESP, was able to inhibit both adhesion and spreading of EC. At three hours, EC attachment and spreading were rapid and comparable on fibronectin and PVAm[Pep (100%)], rounded on PVAm[Pep (0%)], and intermediate on PVAm[Pep (25%)], (PVAm[Pep (50%)], and PVAm[Pep (75%)], with increasing peptide ratio favoring more spreading, although all the substrates had similar hydrophilicity. Cells that spread well on fibronectin and PVAm[Pep (100%)] had sharp spikes of vinculin localized at the termination point of actin stress fibers. Formation of stress fibers and focal adhesions on other substrates were correlated with spreading pattern of EC and the peptide content. EC seeded on fibronectin expressed alpha5beta1 integrins all along the stress fibers and throughout the entire cytoskeleton, but this distribution pattern was less prominent on PVAm[Pep (100%)]. However, expression and distribution of vitronectin receptors (alpha(v)beta3) were similar on both fibronectin and PVAm[Pep (100%)], suggesting a strong cell adhesion on PVAm[Pep (100%)]. Viability of EC was also comparable on both fibronectin and PVAm[Pep (100%)] at 24 h. Substrates with high proportion of dextran limited cell adhesion, probably by decreasing protein adsorption. These results suggest that it may be possible to engineer substrates that promote cell adhesion in a receptor-dependent manner while blocking nonspecific protein adsorption, which may have potential as interface materials for prostheses used in cardiovascular system.     

Susceptibility of bacterial, eukaryotic and artificial membranes to the disruptive action of the cationic peptides Pep 5 and nisin

Abstract The cationic bactericidal peptides Pep 5 and nisin render membranes permeable to low- M _r compounds. All Gram-positive bacteria treated with these peptides showed an immediate efflux of entrapped radioactive markers. The uptake of α-[¹⁴C]methylglucoside by the phosphoenolpyruvate-dependent phosphotransferase system was stimulated by Pep 5, supporting previous results that pep 5 abolishes the membrane potential. Oxygen consumption was inhibited, presumably due to lack of ADP. Escherichia coli became sensitive to Pep 5 and nisin when the outer membrane was bypassed by osmotic shock or by formation of cytoplasmic membrane vesicles. In contrast, Mycoplasma cells and erythrocytes were unaffected by Pep 5 and nisin in concentrations up to 1 mM. Human lung fibroblasts released only small amounts of ATP when treated with Pep 5 and nisin in μM concentrations. Eukaryotic and Mycoplasma cells were disrupted more effectively by the bee venom peptide melittin, which displays overall structural similarities to Pep 5 and nisin. Various artificial membranes were not affected by Pep 5, nisin, or melittin.     

Enhanced detection of human immunodeficiency virus type 1 (HIV-1) Nef-specific T cells recognizing multiple variants in early HIV-1 infection

A human immunodeficiency virus (HIV)-preventive vaccine will likely need to induce broad immunity that can recognize antigens expressed within circulating strains. To understand the potentially relevant responses that T-cell based vaccines should elicit, we examined the ability of T cells from early infected persons to recognize a broad spectrum of potential T-cell epitopes (PTE) expressed by the products encoded by the HIV type 1 (HIV-1) nef gene, which is commonly included in candidate vaccines. T cells were evaluated for gamma interferon (IFN-gamma) secretion using two peptide panels: subtype B consensus (CON) peptides and a novel peptide panel providing 70% coverage of PTE in subtype B HIV-1 Nef. Eighteen of 23 subjects' T cells recognized HIV-1 Nef. In one subject, Nef-specific T cells were detected with the PTE but not with the CON peptides. The greatest frequency of responses spanned Nef amino acids 65 to 103 and 113 to 147, with multiple epitope variants being recognized. Detection of both the epitope domain number and the response magnitude was enhanced using the PTE peptides. On average, we detected 2.7 epitope domains with the PTE peptides versus 1.7 domains with the CON peptides (P = 0.0034). The average response magnitude was 2,169 spot-forming cells (SFC)/10(6) peripheral blood mononuclear cells (PBMC) with the PTE peptides versus 1,010 SFC/10(6) PBMC with CON peptides (P = 0.0046). During early HIV-1 infection, Nef-specific T cells capable of recognizing multiple variants are commonly induced, and these responses are readily detected with the PTE peptide panel. Our findings suggest that Nef responses induced by a given vaccine strain before HIV-1 exposure may be sufficiently broad to recognize most variants within subtype B HIV-1.     

Modeling GPCR active state conformations: the β(2)-adrenergic receptor

The recent publication of several G protein-coupled receptor (GPCR) structures has increased the information available for homology modeling inactive class A GPCRs. Moreover, the opsin crystal structure shows some active features. We have therefore combined information from these two sources to generate an extensively validated model of the active conformation of the β(2)-adrenergic receptor. Experimental information on fully active GPCRs from zinc binding studies, site-directed spin labeling, and other spectroscopic techniques has been used in molecular dynamics simulations. The observed conformational changes reside mainly in transmembrane helix 6 (TM6), with additional small but significant changes in TM5 and TM7. The active model has been validated by manual docking and is in agreement with a large amount of experimental work, including site-directed mutagenesis information. Virtual screening experiments show that the models are selective for β-adrenergic agonists over other GPCR ligands, for (R)- over (S)-β-hydroxy agonists and for β(2)-selective agonists over β(1)-selective agonists. The virtual screens reproduce interactions similar to those generated by manual docking. The C-terminal peptide from a model of the stimulatory G protein, readily docks into the active model in a similar manner to which the C-terminal peptide from transducin, docks into opsin, as shown in a recent opsin crystal structure. This GPCR-G protein model has been used to explain site-directed mutagenesis data on activation. The agreement with experiment suggests a robust model of an active state of the β(2)-adrenergic receptor has been produced. The methodology used here should be transferable to modeling the active state of other GPCRs.     

Hepta and octapeptide agonists of protease-activated receptor 2.

Protease-activated receptor 2 (PAR(2)) is a G protein-coupled cell surface receptor for trypsin-like enzymes. Proteolytic cleavage at a specific site in the extracellular N-terminus exposes a receptor-activating sequence, the 'tethered ligand', which binds intramolecularly to initiate receptor signalling. Peptide or small molecule agonists for PAR(2), devoid of the non-specific and proteolytic effects of enzyme activators, may be promising therapeutic agents for proliferative and inflammatory diseases reportedly mediated by PAR(2). Synthetic hexapeptides that correspond to the native tethered ligand of human or rodent PAR(2) (SLIGKV and SLIGRL, respectively) can activate the receptor independently of proteolytic cleavage; however, known peptide agonists have much lower potency compared to protease-mediated activation. Here, we investigated the agonist activity of 94 hepta and octapeptide derivatives of the human and rodent PAR(2)-tethered ligand sequences in human airway epithelial (A549) cells which endogenously express PAR(2). Thirty synthetic peptides were found to be as potent as or more potent than SLIGRL on the basis of intracellular Ca(2+) responses. The more active peptide agonists were also examined for agonist cross-reactivity at PAR(1) in Chinese Hamster Ovary (CHO) cells that endogenously express functional PAR(1) but not PAR(2). Two potent and PAR(2)-selective agonists were further examined for their capacity to relax phenylephrine-contracted rat aortic rings. Our findings reveal an important role for carboxyl extensions to native PAR(2) activating peptides in potentiating agonist activity.     

Phage display biopanning identifies the translation initiation and elongation factors (IF1α-3 and eIF-3) as components of Hsp70–peptide complexes in breast tumour cells

The heat shock protein, HSP70, is over-expressed in many tumours and acts at the crossroads of key intracellular processes in its role as a molecular chaperone. HSP70 associates with a vast array of peptides, some of which are antigenic and can mount adaptive immune responses against the tumour from which they are derived. The pool of peptides associated with HSP70 represents a unique barcode of protein metabolism in tumour cells. With a view to identifying unique protein targets that may be developed as tumour biomarkers, we used purified HSP70 and its associated peptide pool (HSP70–peptide complexes, HSP70-PCs) from different human breast tumour cell lines as targets for phage display biopanning. Our results show that HSP70-PCs from each cell line interact with unique sets of peptides within the phage display library. One of the peptides, termed IST, enriched in the biopanning process, was used in a ‘pull-down’ assay to identify the original protein from which the HSP70-associated peptides may have been derived. The eukaryotic translation initiation factor 3 (eIF-3), a member of the elongation factor EF1α family, and the HSP GRP78, were pulled down by the IST peptide. All of these proteins are known to be up-regulated in cancer cells. Immunohistochemical staining of tumour tissue microarrays showed that the peptide co-localised with HSP70 in breast tumour tissue. The data indicate that the reservoir of peptides associated with HSP70 can act as a unique indicator of cellular protein activity and a novel source of potential tumour biomarkers.     

The complete amino acid sequence of a biologically active 197-residue fragment of M protein isolated from type 5 group A streptococci

The complete amino acid sequence of a peptic fragment (Pep M5) of the group A streptococcal type 5 M protein, the antiphagocytic cell surface molecule of the bacteria, is described. This fragment, comprising nearly half of the native M molecule, is biologically active in that it has the ability to interact with opsonic antibodies as well as to evoke such an antibody response in rabbits. The sequence of Pep M5 was determined by automated Edman degradations of the uncleaved molecule and its enzymatically derived peptides. The primary peptides for Edman degradation were the arginine peptides obtained by tryptic digestion. The tryptic cleavage of Pep M5 was limited to the arginyl peptide bonds by derivatizing the epsilon-amino groups of lysine residues by reductive dihydroxypropylation. The overlapping peptides were generated by digestion of the unmodified Pep M5 with chymotrypsin, V8 protease, and subtilisin. The sequence thus established for the Pep M5 molecule consists of a total of 197 residues (Mr = 22,705). The Pep M5 protein contains some identical, or nearly so, repeating sequences: four 7-residue segments and two 10-residue segments. However, extensive sequence repeats of the kind previously reported within the partial sequence of another M protein serotype, namely Pep M24, were absent. The Pep M5 sequence is distinct from, but exhibits some homology with, the partial sequences of two other M protein serotypes, namely, Pep M6 and Pep M24. Furthermore, the 7-residue periodicity of the nonpolar and charged residues, an alpha-helical coiled-coil structural characteristic that was previously observed within the partial sequences of M proteins, was found to extend over a significant part of the Pep M5 sequence. The implication of these results to the function and immunological diversity in M proteins is discussed.     

Influence of the staphylococcinlike peptide Pep 5 on membrane potential of bacterial cells and cytoplasmic membrane vesicles.

The staphylococcinlike peptide Pep 5 rapidly abolished the membrane potential of bacterial cells; active transport of amino acids by cytoplasmic membrane vesicles was inhibited and preaccumulated amino acids were released upon the addition of Pep 5. Artificial asolectin vesicles were not impaired by the peptide. It is concluded that the cytoplasmic membrane is the primary target of Pep 5.     

Phylloxin, a novel peptide antibiotic of the dermaseptin family of antimicrobial/opioid peptide precursors.

A novel family of peptide precursors that have very similar N-terminal preprosequences followed by markedly different C-terminal domains has been identified in the skin of hylid frogs belonging to the genus Phyllomedusinae. Biologically active peptides derived from the variable domains include the dermaseptins, 28-34-residue peptides that have a broad-spectrum microbicidal activity, and dermorphin and the deltorphins, D-amino acid containing heptapeptides that are very potent agonists for the micro-opioid and delta-opioid receptors, respectively. This report describes the isolation, synthesis and cloning of phylloxin, a prototypical member of a novel family of antimicrobial peptides derived from the processing of a dermaseptin/dermorphin-like precursor. The structure of phylloxin (GWMSKIASGIGTFLSGIQQ amide) shows no homology to the dermaseptins, but bears some resemblance to the levitide-precursor fragment and the xenopsin-precursor fragment, two antimicrobial peptides isolated from the skin of an evolutionarily distant frog species, Xenopus laevis. Circular dichroism spectra of phylloxin in low polarity medium, which mimics the lipophilicity of the membrane of target microorganisms, indicated 60-70% alpha-helical conformation, and predictions of secondary structure suggested that the peptide can be configured as an amphipathic helix spanning residues 1-19. Phylloxin is an addition to the structurally and functionally diverse peptide families encoded by the rapidly evolving C-terminal domains of the dermorphin/dermaseptin group of precursors.