Helicobacter Pylori Ribosomal Protein L1 Research Articles

Anticancer activity of Helicobacter pylori ribosomal protein (HPRP) with iRGD in treatment of colon cancer.

As the conventional therapeutic approaches were not completely successful in the treatment of colon cancer, there is still a need for finding the most efficient therapeutic agents. Here we investigated the anticancer activity of HPRP-A1 that was derived from the N-terminal region of Helicobacter pylori ribosomal protein L1 (RpL1) alone or in combination with tumor-homing peptide iRGD and 5-Fluorouracil (5FU) on colon cancer cell lines (CT26 and HT29) and isograft models of colon cancer. We assessed the tumor growth inhibitory activity of HPRP-A1 with or without iRGD and 5FU on colon cancer in vitro and in vivo. In the in vitro part, we investigate the effect of HPRP-A1 alone and in combination with iRGD/5FU. Our results demonstrated that co-administration of HPRP-A1 with iRGD increased the apoptosis, while these two peptides in combination with 5FU increased the intracellular level of p53 that upregulate the pro-apoptotic gene BAX and downregulate the anti-apoptotic gene BCL2. HPRP-A1 blocks the cell cycle progression in G0/G1. Co-administration of two peptides significantly reduced the size and weight of the tumors, while the group that received 5FU in combination with the peptides increased the necrotic and decrease the fibrotic area significantly in the tumor tissues, which also disrupt the oxidant/antioxidant balance. Our results indicated that HPRP-A1 could be considered an effective agent toward colon cancer in vitro and in vivo with the ability to enhance the effects of conventional chemotherapy agent 5FU.

All d-Lysine Analogues of the Antimicrobial Peptide HPA3NT3-A2 Increased Serum Stability and without Drug Resistance

Novel antibiotic drugs are urgently needed because of the increase in drug-resistant bacteria. The use of antimicrobial peptides has been suggested to replace antibiotics as they have strong antimicrobial activity and can be extracted from living organisms such as insects, marine organisms, and mammals. HPA3NT3-A2 ([Ala1,8] HPA3NT3) is an antimicrobial peptide that is an analogue of the HP (2–20) peptide derived from Helicobacter pylori ribosomal protein L1. Although this peptide was shown to have strong antimicrobial activity against drug-resistant bacteria, it also showed lower toxicity against sheep red blood cells (RBCs) and HaCaT cells compared to HPA3NT3. The l-Lys residues of HPA3NT3-A2 was substituted with d-Lys residues (HPA3NT3-A2D; [d-Lys2,5,6,9,10,15] HPA3NT3-A2) to prevent the cleavage of peptide bonds by proteolytic enzymes under physiological conditions. This peptide showed an increased half-life and maintained its antimicrobial activity in the serum against Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) (pathogen). Furthermore, the antimicrobial activity of HPA3NT3-A2D was not significantly affected in the presence of mono- or divalent ions (Na+, Mg2+, Ca2+). Finally, l- or d-HPA3NT3-A2 peptides exhibited the strongest antimicrobial activity against antibiotic-resistant bacteria and failed to induce resistance in Staphylococcus aureus after 12 passages.

A helix-PXXP-helix peptide with antibacterial activity without cytotoxicity against MDRPA-infected mice

In response to the growing problem of multidrug-resistant pathogenic microbes, much attention is being paid to naturally occurring and synthetic antimicrobial peptides (AMPs) and the effects of their structural modification. Among these modifications, amino acid substitution is a simple approach to enhancing biological activity and reducing cytotoxicity. An earlier study indicated that HPA3, an analog of HP (2-20) derived from the N-terminus of Helicobacter pylori ribosomal protein L1, forms large pores and shows considerable cytotoxicity. However, HPA3P, in which a proline (Pro) is substituted for glutamic acid (Glu) at position 9 of HPA3, shows markedly less cytotoxicity. This may be attributable to the presence of a Pro-kink into middle of the HPA3P structure within the membrane environment. Unfortunately, HPA3P is not an effective antibacterial agent in vivo. We therefore designed a helix-PXXP-helix structure (HPA3P2), in which Pro was substituted for the Glu and phenylalanine (Phe) at positions 9 and 12 of HPA3, yielding a molecule with a flexible central hinge. As compared to HPA3P, HPA3P3 exhibited dramatically increased antibacterial activity in vivo. ICR mice infected with clinically isolated multidrug-resistant Pseudomonas aeruginosa showed 100% survival when administered one 0.5-mg/kg dose of HPA3P2 or three 0.1-mg/kg doses of HPA3P2. Moreover, in a mouse model of septic shock induced by P. aeruginosa LPS, HPA3P2 reduced production of pro-inflammatory mediators and correspondingly reduced lung (alveolar) and liver tissue damage. The changes in HPA3 behavior with the introduction of Pro likely reflects alterations of the mechanism of action: i) HPA3 forms pores in the bacterial cell membranes, ii) HPA3P permeates the cell membranes and binds to intracellular RNA and DNA, and iii) HPA3P2 acts on the outer cellular membrane component LPS. Collectively, these results suggest HPA3P2 has the potential to be an effective antibiotic for use against multidrug-resistant bacterial strains.

A proline-hinge alters the characteristics of the amphipathic α-helical AMPs.

HP (2–20) is a 19-aa, amphipathic, α-helical peptide with antimicrobial properties that was derived from the N-terminus of Helicobacter pylori ribosomal protein L1. We previously showed that increasing the net hydrophobicity of HP (2–20) by substituting Trp for Gln17 and Asp19 (Anal 3) increased the peptide's antimicrobial activity. In hydrophobic medium, Anal 3 forms an amphipathic structure consisting of an N-terminal random coil region (residues 2–5) and an extended helical region (residues 6–20). To investigate the structure-activity relationship of Anal 3, we substituted Pro for Glu9 (Anal 3-Pro) and then examined the new peptide's three-dimensional structure, antimicrobial activity and mechanism of action. Anal 3-Pro had an α-helical structure in the presence of trifluoroethanol (TFE) and sodium dodecyl sulfate (SDS). NMR spectroscopic analysis of Anal 3-Pro's tertiary structure in SDS micelles confirmed that the kink potential introduced by Pro10 was responsible for the helix distortion. We also found that Anal 3-Pro exhibited about 4 times greater antimicrobial activity than Anal 3. Fluorescence activated flow cytometry and confocal fluorescence microscopy showed that incorporating a Pro-hinge into Anal 3 markedly reduced its membrane permeability so that it accumulated in the cytoplasm without remaining in the cell membrane. To investigate the translocation mechanism, we assessed its ability to release of FITC-dextran. The result showed Anal 3-Pro created a pore <1.8 nm in diameter, which is similar to buforin II. Notably, scanning electron microscopic observation of Candida albicans revealed that Anal 3-Pro and buforin II exert similar effects on cell membranes, whereas magainin 2 exerts a different, more damaging, effect. In addition, Anal 3-Pro assumed a helix-hinge-helix structure in the presence of biological membranes and formed micropores in both bacterial and fungal membranes, through which it entered the cytoplasm and tightly bound to DNA. These results indicate that the bending region of Anal 3- Pro peptide is prerequisite for effective antibiotic activity and may facilitate easy penetration of the lipid bilayers of the cell membrane.

Antimicrobial HPA3NT3 peptide analogs: Placement of aromatic rings and positive charges are key determinants for cell selectivity and mechanism of action

In an earlier study, we determined that HP(2‐20) (residues 2‐20 of parental HP derived from the N-terminus of the Helicobacter pylori ribosomal protein L1) and its analog, HPA3NT3, had potent antimicrobial effects. However, HPA3NT3 also showed undesirable cytotoxicity against HaCaT cells. In the present study, we designed peptide analogs including HPA3NT3-F1A (‐F1A), HPA3NT3-F8A (‐F8A), HPA3NT3-F1AF8A (‐F1AF8A), HPA3NT3-A1 (‐A1) and HPA3NT3-A2 (‐A2) in an effort to investigate the effects of amino acid substitutions in reducing their hydrophobicity or increasing their cationicity, and any resulting effects on their selectivity in their interactions with human cells and pathogens, as well as their mechanism of antimicrobial action. With the exception of HPA3NT3-A1, all of these peptides showed potent antimicrobial activity. Moreover, substitution of Ala for Phe at positions 1 and/or 8 of the HPA3NT3 peptides (‐F1A, -F8A and -F1AF8A) dramatically reduced their cytotoxicity. Thus the cytotoxicity of HPA3NT3 appears to be related to its Phe residues (positions 1 and 8), which strongly interact with sphingomyelin in the mammalian cell membrane. HPA3NT3 exerted its bactericidal effects through membrane permeabilization mediated by pore formation. In contrast, fluorescent dye leakage and nucleic acid gel retardation assays showed that ‐A2 acted by penetrating into the cytoplasm, where it bound to nucleic acids and inhibited protein synthesis. Notably, Staphylococcus aureus did not develop resistance to -A2 as it did with rifampin. These results suggest that the -A2 peptide could potentially serve as an effective antibiotic agent against multidrug-resistant bacterial strains.

Novel Short AMP: Design and Activity Study

In a previous study, we reported that truncation of HP (2-20) (derived from the N-terminal region of Helicobacter pylori Ribosomal Protein L1 (RPL1)) at the N- (residues 2-3) and C-terminal (residues 17-20) truncated fragments to give HP (4-16) induces increased antibiotic activity against several bacterial strains without hemolysis. In this study, to develop novel short antibiotic peptides useful as therapeutic drugs, an analogue was designed to possess increased hydrophobicity by Trp substitution in position 2 region of HP (4-16). Synthetic HP (4-16)-W showed an enhanced antimicrobial and antitumor activity. The antimicrobial activity of this peptide and others was measured by their growth inhibitory effect upon S. aureus, B. subtilis, S. epidermidis, E. coli, S. typimurium, P. aeruginosa, C. albicans, T. beigelii and S. cerevisiae. None of the peptides exhibited hemolytic activity against human erythrocyte cells except melittin as a positive control. Its antibiotic activity suggests that HP (4-16)-W is an excellent candidate as a lead compound for the development of novel antibiotic agents.

Effect of acyl chain structure and bilayer phase state on binding and penetration of a supported lipid bilayer by HPA3

The effect of acyl chain structure and bilayer phase state on binding and penetration by the peptide HPA3 was studied using dual polarisation interferometry. This peptide is an analogue of Hp(2-20) derived from the N-terminus of Helicobacter pylori ribosomal protein L1 (RpL1) which has been shown to have antimicrobial and cell-penetrating properties. The binding of HPA3 to zwitterionic 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) or 1-palmitolyl-2-oleyl-sn-glycero-3-phosphocholine (POPC) and negatively charged membranes composed of DMPC and 1,2-dimyristoyl-sn-glycero-3-(phosphor-rac-(1-glycerol)) (DMPG) or POPC and 1-palmitolyl-2-oleyl-sn-glycero-3-(phosphor-rac-(1-glycerol)) (POPG) was determined using dual polarisation interferometry (DPI). Mass and birefringence were measured in real time, enabling the creation of birefringence-mass plots for detailed analysis of the changes in lipid bilayer order during the peptide-binding process. HPA3 bound to all four lipids and the binding progressed as a single phase for the saturated gel phase bilayers DMPC and DMPC-DMPG. However, the binding process involved two or more phases, with penetration of the unsaturated fluid phase POPC and POPC-POPG bilayers. Structural changes in the saturated bilayer were partially reversible whereas binding to the unsaturated bilayer resulted in irreversible changes in membrane structure. These results demonstrate that more disordered unsaturated bilayers are more susceptible to further disorganisation and have a lower capacity to recover from peptide-induced structural changes than saturated ordered bilayers. In addition, this study further establishes DPI as powerful tool for analysis of multiphase peptide-insertion processes associated with complex structural changes in the liquid-crystalline membrane.

The membrane insertion of helical antimicrobial peptides from the N-terminus of Helicobacter pylori ribosomal protein L1

The interaction of two helical antimicrobial peptides, HPA3 and HPA3P with planar supported lipid membranes was quantitatively analysed using two complementary optical biosensors. The peptides are analogues of Hp(2-20) derived from the N-terminus of Helicobacter pylori ribosomal protein L1 (RpL1). The binding of these two peptide analogues to zwitterionic dimyristoyl-phosphatidylcholine (DMPC) and negatively charged membranes composed of DMPC/dimyristoylphosphatidylglycerol (DMPG) (4:1) was determined using surface plasmon resonance (SPR) and dual polarisation interferometry (DPI). Using SPR analysis, it was shown that the proline substitution in HPA3P resulted in much lower binding for both zwitterionic and anionic membranes than HPA3. Structural changes in the planar DMPC and DMPC/DMPG (4:1) bilayers induced by the binding of both Hp(2-20) analogues were then resolved in real-time with DPI. The overall process of peptide-induced changes in membrane structure was analysed by the real-time changes in bound peptide mass as a function of bilayer birefringence. The insertion of both HPA3 and HPA3P into the supported lipid bilayers resulted in a decrease in birefringence with increasing amounts of bound peptide which reflects a decrease in the order of the bilayer. The binding of HPA3 to each membrane was associated with a higher level of bound peptide and greater membrane lipid disordering and a faster and higher degree of insertion into the membrane than HPA3P. Furthermore, the binding of both HPA3 and HPA3P to negatively charged DMPC/DMPG bilayers also leads to a greater disruption of the lipid ordering. These results demonstrate the geometrical changes in the membrane upon peptide insertion and the extent of membrane structural changes can be obtained quantitatively. Moreover, monitoring the effect of peptides on a structurally characterised bilayer has provided further insight into the role of membrane structure changes in the molecular basis of peptide selectivity and activity and may assist in defining the mode of antimicrobial action.

Effects of Substitutions of Gln16 and Asp18 with Phe or Tyr in HP(2-20) on its Structure and Antimicrobial Activity

HP (2-20), a 19-residue peptide derived from the N-terminus of Helicobacter pylori Ribosomal Protein L1, has antimicrobial activity but is not cytotoxic to human erythrocytes. Previously, we have synthesized several analogue peptides to investigate the effects of substitutions on the structure and antimicrobial activity. Substitution of and with Trp (Anal 3) caused a dramatic increase in bacterial and fungal lytic activities. In this study, analogue peptides were synthesized to investigate the effects of substitution of Gin and Asp with Phe (Anal 6) or Tyr (Anal 7) in HP (2-20) on its structure and antimicrobial activity. Substitution of Gin and Asp with hydrophobic aromatic residues at position 16 and 18 of HP (2-20) caused increase in antibiotic activity without hemolytic effect. Substitution of Gin and Asp with Trp and Try increased antibiotic activity of HP (220) twice more compared to substitution with Phe. The tertiary structures of Anal 6 and Anal 7 in SDS micelles has been investigated using NMR spectroscopy. The structures revealed that substitutions of the aromatic residues at C-terminus resulted in longer and well defined alpha-helix and improved their antibacterial activities

Amphipathic α-helical peptide, HP (2–20), and its analogues derived from Helicobacter pylori: Pore formation mechanism in various lipid compositions

In a previous study, we determined that HP(2–20) (residues 2–20 of parental HP derived from the N-terminus of Helicobacter pylori Ribosomal Protein L1) and its analogue, HPA3, exhibit broad-spectrum antimicrobial activity. The primary objective of the present study was to gain insight into the relevant mechanisms of action using analogues of HP(2–20) together with model liposomes of various lipid compositions and electron microscopy. We determined that these analogues, HPA3 and HPA3NT3, exert potent antibacterial effects in low-salt buffer and antifungal activity against chitin-containing fungi, while having little or no hemolytic activity or cytotoxicity against mammalian cell lines. Our examination of the interaction of HP(2–20) and its analogues with liposomes showed that the peptides disturb both neutral and negatively-charged membranes, as demonstrated by the release of encapsulated fluorescent markers. The release of fluorescent markers induced by HP(2–20) and its analogues was inversely related to marker size. The pore created by HP(2–20) shows that the radius is approximately 1.8 nm, whereas HPA3, HPA3NT3, and melittin have apparent radii between 3.3 and 4.8 nm. Finally, as shown by electron microscopy, the liposomes and various microbial cells treated with HPA3 and HPA3NT3 showed oligomerization and blebbing similar to that seen with melittin, while HP(2–20) exhibited flabbiness. These results suggest that HP(2–20) may exert its antibiotic effects through a small pore (about 1.8 nm), whereas HPA3 and HPA3NT3 formed pores of a size consistent with those formed by melittin.

Structure‐activity relationship of HP (2–20) analog peptide: Enhanced antimicrobial activity by N‐terminal random coil region deletion

HP (2-20) (AKKVFKRLEKLFSKIQNDK) is a 19-aa antimicrobial peptide derived from N-terminus of Helicobacter pylori Ribosomal protein L1 (RpL1). In the previous study, several analogs with amino acid substitutions were designed to increase or decrease only the net hydrophobicity. In particular, substitutions of Gln(16) and Asp(18) with Trp (Anal 3) for hydrophobic amino acid caused a dramatic increase in antibiotic activity without a hemolytic effect. HP-A3 is a potent antimicrobial peptide that forms, in a hydrophobic medium, an amphipathic structure consisting of an N-terminal random coil region (residues 2-5) and extended C-terminal regular alpha-helical region (residues 6-20). To obtain the short and potent alpha-helical antimicrobial peptide, we synthesized a N-terminal random coil deleted HP-A3 (A3-NT) and examined their antimicrobial activity and mechanism of action. The resulting 15mer peptide showed increased antibacterial and antifungal activity to 2- and 4-fold, respectively, without hemolysis. Confocal fluorescence microscopy studies showed that A3-NT was accumulated in the plasma membrane. Flow cytometric analysis revealed that A3-NT acted in salt- and energy-independent manner. Furthermore, A3-NT causes significant morphological alterations of the bacterial surfaces as shown by scanning electron microscopy. Circular dichroism (CD) analysis revealed that A3-NT showed higher alpha-helical contents than the HP-A3 peptide in 50% TFE solution. Therefore, the cell-lytic efficiency of HP-A3, which depended on the alpha-helical content of peptide, correlated linearly with their antimicrobial potency.

Interactions between the plasma membrane and the antimicrobial peptide HP (2-20) and its analogues derived from Helicobacter pylori

HP (2-20), a 19-residue peptide derived from the N-terminus of Helicobacter pylori ribosomal protein L1, has antimicrobial activity but is not cytotoxic to human erythrocytes. We synthesized several peptide analogues to investigate the effects of substitutions on structure and antimicrobial activity. Replacement of Gln16 and Asp18 with tryptophan [anal-3 (analogue-3)] caused a dramatic increase in lytic activities against bacteria and fungi. By contrast, a decrease in amphiphilicity caused by replacement of Phe5 or Leu11 with serine was accompanied by a reduction in antimicrobial activity. Analysis of the tertiary structures of the peptides in SDS micelles by NMR spectroscopy revealed that they have a well-defined a-helical structure. Among the analogues, anal-3 has the longest a-helix, from Val4 to Trp18. The enhanced hydrophobicity and increased a-helicity results in enhanced antimicrobial activity in anal-3 without an increase in haemolytic activity. Fluorescence experiments proved that the bacterial-cell selectivity of the anal-3 peptide is due to its high binding affinity for negatively charged phospholipids in bacterial cells. Results showing the effect of spin-labels on the NMR spectra indicated that the side chains in the hydrophobic phase of the amphiphilic a-helix are buried on the surface of the micelle and the tryptophan indole ring is anchored in the membrane surface. Because anal-3 shows high selectivity towards bacterial and fungal cells, it may provide an avenue for the development of new antibiotics.

Antibiotic Activity of Antimicrobial Peptide Against Pseudomonads Isolated from a Patient with Gallstones

We investigated the in vitro antibiotic activity of the 19-amino acid antimicrobial peptide HP (2-20), derived from the N-terminus of Helicobacter pylori Ribosomal Protein L1 (RPL1), against antibiotic susceptible and resistant pathogens from a patient with gallstones. HP (2-20) was active against antibiotic-susceptible and antibiotic-resistant clinical isolates of pathogens from a patient with gallstones, but this peptide showed no hemolytic activity against normal human erythrocytes. HP (2-20) acted synergistically with ciprofloxacin against pathogenic bacteria. Fluorescence activated flow cytometry revealed that the effect of HP (2-20) was dependent on energy and salt concentration. In addition, scanning electron microscopy showed that HP (2-20) caused significant morphological alterations to the cell surface of pathogens. Using 16S rDNA sequences, we found that isolates from bile were 100% homologous to Pseudomonas aeruginosa. These findings suggest that HP (2-20) may be useful clinically as an antibiotic against acquired pathogens from patients with gallstones and against pathogens resistant to other antibiotics.

Effects of N- and C-terminal truncation of HP (2-20) from Helicobacter pylori ribosomal protein L1 (RPL1) on its anti-microbial activity

HP (2-20) [derived from the N-terminal region of Helicobacter pylori Ribosomal Protein L1 (RPL1)], a 19-mer peptide, possesses broad-spectrum anti-microbial activity. As the N- (residues 2-3) and C-terminal (residues 14-20) residues can be deleted without affecting antimicrobial activity, we have now determined the minimum chain length necessary for the retention of antimicrobial activity, and its mode of action. The N- (residues 2-3) and C-terminal (residues 17-20) truncated fragments [HP (4-16)] induce increased antibiotic activity against several bacterial strains without hemolysis. Flow cytometric analysis, scanning electron microscopy and fluorescence confocal microscopy revealed that HP (4-16) acted rapidly on the plasma membranes of the fungal cells in a salt- and energy-independent manner.

Antibiotic activity and synergistic effect of antimicrobial peptide against pathogens from a patient with gallstones

HP (2–20) is a peptide derived from the N-terminus of Helicobacter pylori ribosomal protein L1 that has been shown to have antimicrobial activity against various species of bacteria. When we tested the effects of HP (2–20), we found that this peptide displayed strong activity against pathogens from a patient with gallstones, but it did not have hemolytic activity against human erythrocytes. We also found that HP (2–20) had potent activity against cefazolin sodium-resistant bacterial cell lines, and that HP (2–20) and cefazolin sodium had synergistic effects against cell lines resistant to the latter. To investigate the mechanism of action of HP (2–20), we performed fluorescence activated flow cytometry using pathogens from the patient with gallstones. As determined by propidium iodide (PI) staining, pathogenic bacteria treated with HP (2–20) showed higher fluorescence intensity than untreated cells, similar to melittin-treated cells, and that HP (2–20) acted in an energy- and salt-dependent manner. Scanning electron microscopy showed that HP (2–20) caused significant morphological alterations in the cell surface of pathogens from the patient with gallstones. By determining their 16S rDNA sequences, we found that both the pathogens from the patient with gallstones and the cefazolin sodium-resistant cell lines showed 100% homology with sequences from Pseudomonas aeruginosa. Taken together, these results suggest that HP (2–20) has antibiotic activity and that it may be used as a lead drug for the treatment of acquired pathogens from patients with gallstones and antibiotic-resistant cell lines.

Antibacterial synergism of novel antibiotic peptides with chloramphenicol

HP (2–20) is an antimicrobial sequence derived from the N-terminus of Helicobacter pylori ribosomal protein L1. We previously tested whether several analogues of HP (2–20), with amino acid substitutions that increased or decreased net hydrophobicity, could be useful as therapeutic agents. In the present study, we show that substituting Gln and Asp for Trp at positions 17 and 19, respectively, of HP (2–20) (peptide A3) had potent antibacterial activity in minimal inhibition concentration and minimal bactericidal concentration without having hemolytic activity. In contrast, when we decreased hydrophobicity by substituting Leu or Phe for Ser at positions 12 and 19, respectively, of HP (2–20) (Anal 4, Anal 5), there was no significant effect on antibacterial activity. We found that A3 acted synergistically with chloramphenicol against bacterial cells. Fluorescence activated flow cytometry showed that A3-treated cells had higher fluorescence intensity than untreated cells, similar to that of melittin-treated cells. Furthermore, A3 caused significant morphological alterations of Staphylococcus aureus and Pseudomonas aeruginosa, as shown by scanning electron microscopy. Our results suggest that peptide A3 may be useful for the design of novel antibiotic peptides that possess high bacterial cell selectively and synergistic effects with conventional antibiotic agents but lack hemolytic activity.

Antinematodal activity and the mechanism of the antimicrobial peptide, HP (2-20), against Caenorhabditis elegans.

The peptide HP (2-20), derived from the N-terminal sequence of Helicobacter pylori ribosomal protein L1 (RPL1), has a nematicidal activity against eggs and worms of Caenorhabditis elegans. Eggs treated with HP (2-20) (69%) has a higher fluorescence intensity with propidium iodide staining, which was similar to that of melittin (82%) but higher than untreated cells (5.7%). Confocal microscopy showed that the peptides were located in the shell of the eggs and the inner and outer surfaces of the worms. HP (2-20) therefore may exert its antinematodal activity by disrupting the structure of the egg's shell and the cell membrane via pore formation or by direct interaction with the lipid bilayers in a detergent-like manner.

HP(2–9)‐magainin 2(1–12), a synthetic hybrid peptide, exerts its antifungal effect onCandida albicansby damaging the plasma membrane

In our previous study, HP(2-9)-MA(1-12), HP-MA for short, a hybrid peptide incorporating residues 2-9 of Helicobacter pylori ribosomal protein L1 (HP) and residues 1-12 of magainin 2 (MA) was shown to have strong antibacterial activity. In this study the antifungal activity of HP-MA was evaluated using various fungi, and it was shown that the activity was increased when compared with the parent peptides. In order to investigate the fungicidal mechanism(s) of HP-MA its action against fungal cell membranes was examined by the potassium-release test, which showed that HP-MA caused an increase in the amount of K+ released from the cells. Furthermore, HP-MA induced significant morphological changes. These facts suggested that the fungicidal effect of HP-MA involves damaging the fungal cell membranes. CD investigators suggested that the alpha-helical structure of these peptides plays an important role in their antibiotic effect, but that alpha-helicity is less directly correlated with the enhanced antibiotic activity of the hybrid.

Virus-cell fusion inhibitory activity of novel analogue peptides based on the HP (2-20) derived from N-terminus of Helicobacter pylori Ribosomal Protein L1.

HP (2-20) (AKKVFKRLEKLFSKIQNDK) is the antibacterial sequence derived from N-terminus of Helicobacter pylori Ribosomal Protein L1 (RPL1). It has a broad-spectrum microbicidal activity in vitro that is thought to be related to the membrane-disruptive properties of the peptide. Based on the putative membrane-targeted mode of action, we postulated that HP (2-20) might be possessed virus-cell fusion inhibitory activity. To develop the novel virus-cell fusion inhibitory peptides, several analogues with amino acid substitution were designed to increase or decrease only net hydrophobic region. In particular, substitution of Gln and Asp for hydrophobic amino acid, Trp at position 17 and 19 of HP (2-20) (Anal 3) caused a dramatic increase in virus-cell fusion inhibitory activity without hemolytic effect.

HP (2-20) derived from the amino terminal region of helicobacterpylori ribosomal protein L1 exerts its antifungal effects by damaging the plasma membranes of Candida albicans.

The fungicidal effects of the peptide HP (2-20). derived from the N-terminal sequence of Helicobacter pylori ribosomal protein L1 (RPL1). have been investigated. HP (2-20) displays a strong fungicidal activity against various fungi, without haemolytic activity against human erythrocyte cells, and the fungicidal activity is inhibited by Ca2+ and Mg2+ ions. In order to investigate the fungicidal mechanism(s) of HP (2-20). the amount of intracellular trehalose was measured in C. albicans. It was found that the amounts of intracellular trehalose were decreased when HP (2-20) was used. The action of the peptide against fungal cell membranes was further examined by the potassium-release test; HP (2-20) was found to increase the amount of K+ released from the cells. Furthermore, HP (2-20) caused significant morphological changes, as shown by scanning electron microscopy, and by testing the membrane disrupting activity using liposomes (phosphatidyl choline/cholesterol; 10: 1, w/w). Our results suggest that HP (2-20) may exert its antifungal activity by disrupting the structure of cell membranes, via pore formation or direct interaction with the lipid bilayers.