• Bulletin of the Korean Chemical Society
• /
• v.35 no.12
• /
• pp.3632-3636
• /
• 2014

In this study we designed and synthesized several heptapeptides that are enforced to form an amphipathic helix using all-hydrocarbon stapling system and evaluated their antimicrobial and hemolytic activities. The antimicrobial activity showed clear structure-activity relationships, confirming the importance of helicity and amphipathicity. Some stapled heptapeptides displayed a moderate antimicrobial activity along with a low hemolytic activity. To our best knowledge, although not highly potent, these stapled peptides represent the shortest helical amphipathic antimicrobial peptides reported to date. The preliminary data obtained in this work would serve as a good starting point for further developing short analogs of amphipathic helical antimicrobial peptides.

• BMB Reports
• /
• v.43 no.5
• /
• pp.362-368
• /
• 2010

Dermcidin is a human antibiotic peptide that is secreted by the sweat glands and has no homology to other known antimicrobial peptides. As an initial step toward understanding dermcidin's mode of action at bacterial membranes, we used homonuclear and heteronuclear NMR to determine the conformation of the peptide in 50% trifluoroethanol solution. We found that dermcidin adopts a flexible amphipathic $\alpha$-helical structure with a helix-hinge-helix motif, which is a common molecular fold among antimicrobial peptides. Spin-down assays of dermcidin and several related peptides revealed that the affinity with which dermcidin binds to bacterial-mimetic membranes is primarily dependent on its amphipathic $\alpha$-helical structure and its length (>30 residues); its negative net charge and acidic pI have little effect on binding. These findings suggest that the mode of action of dermcidin is similar to that of other membrane-targeting antimicrobial peptides, though the details of its antimicrobial action remain to be determined.

• Bulletin of the Korean Chemical Society
• /
• v.24 no.10
• /
• pp.1478-1484
• /
• 2003

CRAMP was identified from a cDNA clone derived from a mouse femoral marrow cells as a member of cathelicidin-derived antimicrobial peptide. Tertiary structure of CRAMP in TFE/$H_2O$ (1 : 1, v/v) solution has been determined by NMR spectroscopy previously and consists of two amphipathic $\alpha-helices$ from Leu4 to Lys10 and from Gly16 to Leu33. These two helices are connected by a flexible region from Gly11 to Gly16. Analysis of series of fragments composed of various portion of CRAMP revealed that an 18-residue fragment with the sequence from Gly16 to Leu33 (CRAMP-18) was found to retain antibacterial activity without cytotoxicity. The effects of two Phe residues at positions 14 and 15 of CRAMP-18 on structure, antibacterial activity, and interaction with lipid membranes were investigated by $Phe^{14,15}$ ${\rightarrow}$ Ala substitution (CRAMP-18-A) in the present study. Substitution of Phe with Ala in CRAMP-18 caused a significant reduction on antibacterial and membrane-disrupting activities. Tertiary structures of CRAMP-18 in 50% TFE/$H_2O$ (1 : 1, v : v) solution shows amphipathic ${\alpha}$-helix, from $Glu^2{\;}to{\;}Leu^{18}$, while CRAMP-18-A has relatively short amphipathic ${\alpha}$-helix from $Leu^4{\;}to{\;}Ala^{15}$. These results suggest that the hydrophobic property of $Phe^{14}{\;}and{\;}Phe^15$ in CRAMP-18 is essential for its antibacterial activity, ${\alpha}$-helical structure, and interactions with phospholipid membranes.

• Journal of Plant Biology
• /
• v.38 no.4
• /
• pp.359-364
• /
• 1995

As the first step to elucidate the relationship between the structure and function of CTP:phosphocholine cytidylyltransferase (EC 2.7.7.15) in plants, the partial nucleotide sequence of soybean cytidylyltransferase cDNA was determined using a polymerase chain reaction (PCR). Degenerate oligonucleotide primers were synthesized from the conserved region revealed from the rat and yeast cytidylyltransferase DNA sequences. The catalytic domain region showed 78 and 76% homology with the rat and yeast amino acid sequences, respectivly. The hydropathy profile indicated that the C-terminal non-catalytic portion of the protein was very hydrophilic, and in the region between the catalytic domain and the C-terminal region, there was a large amphipathic $\alpha$-helical domain that was believed to bind the membrane surface in the active formation. There are 7 potential sites for phosphorylation by protein kinase C and 4 potential sites for phosphorylation by Ca2+/calmodulin kinase within the determined sequence.

• International Journal of Industrial Entomology and Biomaterials
• /
• v.3 no.2
• /
• pp.163-168
• /
• 2001

We isolated and sequenced a cDNA clone corresponding to apolipophorin-III (apoLp-III) from the fall webworm, Hyphantria cunea. The cDNA for apoLp-III codes fer a 187-residue protein (561 bp) with a predicted molecular mass of 20 kDa. The calculated isoelectric point is 8.76. Multiple alignment analysis of the amino acid sequence revealed that H. cunea apoLp-III is most similar to that of Spodoptera litura (71.5% identity), followed by that of Manduca sexta (69.7% identity). They share five amphipathic $\alpha$-helices that are proposed to play a critical role in the binding of apoLp-III to lipophorin.

• Korean Journal of Microbiology
• /
• v.47 no.4
• /
• pp.328-334
• /
• 2011

Leucine zipper motif consists of multiple periodic leucine residues, which forms amphipathic alpha helix. The hydrophobic nature of leucine zipper motif can dimerize proteins which contain this motif. Leucine zipper motif addition at C-terminus of single-chain Fv (ScFv) antibody induces its dimerization. Since the dimeric ScFv antibody contains two antigen binding sites (bivalency) like Y-shaped complete antibody, it could increase avidity. As a result, it could show higher antigen binding activity than monomeric ScFv antibodies. Based on this concept, monomeric chicken 8C3 ScFv antibody previously developed from chicken hybridoma was dimerized by the addition of leucine zipper motif at C-terminus of ScFv antibody. The dimeric 8C3 ScFv antibody specifically reacted with Eimerian sporozoite which causes Avian Coccidiosis. As expected, dimeric 8C3 ScFv antibody showed 3-folds higher antigen binding activity than monomer due to increased avidity. In addition, protien yields of dimer expression were 2-folds higher than monomer.

• Journal of the Korean Magnetic Resonance Society
• /
• v.19 no.2
• /
• pp.54-60
• /
• 2015

Papiliocin, from the swallowtail butterfly, Papilio xuthus, shows high bacterial cell selectivity against Gram-negative bacteria. Recently, we designed a 22mer analog with N-terminal helix from $Lys^3$ to $Ala^{22}$, PapN. It shows outstanding antimicrobial activity against Gram-negative bacteria with low toxicity against mammalian cells. In this study, we determined the 3-D structure of PapN in 300 mM DPC micelle using NMR spectroscopy and investigated the interactions between PapN and DPC micelles. The results showed that PapN has an amphipathic ${\alpha}$-helical structure from $Lys^3$ to $Lys^{21}$. STD-NMR and DOSY experiment showed that this helix is important in binding to the bacterial cell membrane. Furthermore, we tested antibacterial activities of PapN in the presence of salt for therapeutic application. PapN was calcium- and magnesium-resistant in a physiological condition, especially against Gram-negative bacteria, implying that it can be a potent candidate as peptide antibiotics.

• BMB Reports
• /
• v.32 no.6
• /
• pp.561-566
• /
• 1999

CA(1-8)-ME(1-12) [CA-ME], composed of cecropin A(1-8) and melittin(1-12), is a synthetic antimicrobial peptide having potent antibacterial and antitumor activities with minimal hemolytic activity. In order to investigate the effects of the flexible hinge sequence, Gly-Ile-Gly, of CA-ME on antibiotic activity, CA-ME and three analogues, CA-ME1, CA-ME2, and CA-ME3, were synthesized. The Gly-Ile-Gly sequence of Ca-ME was deleted in CA-ME1 and replaced with Pro and Gly-Pro-Gly in CA-ME2 and CA-ME3, respectively. CA-ME1 and CA-ME3 showed a significant decrease in antitumor activity and phospholipid vesicle-disrupting ability. However, CA-ME2 showed similar antitumor and vesicle-disrupting activities, as compared with CA-ME. These results suggest that the flexibility or ${\beta}$-turn induced by Gly-Ile-Gly or Pro in the central part of CA-ME may be important in the electrostatic interaction of the N-terminus cationic ${\alpha}$-helical region with the cell membrane surface and the hydrophobic interaction of the C-terminus amphipathic ${\alpha}$-helical region with the hydrophobic acyl chains in the cell membrane. CA-ME3 exhibited lower antitumor and vesicle-disrupting activities than CA-ME and CA-ME2. This result suggests that the excessive ${\beta}$-turn structure caused by the Gly-Pro-Gly sequence in CA-ME3 seems to interrupt ion channel/pore formation in the lipid bilayer. We concluded that the appropriate flexibility or bilayer. We concluded that the appropriate flexibility or ${\beta}$-turn structure provided by the central hinge is responsible for the effective antibiotic activity of the antimicrobial peptides with the helix-hinge-helix structure.

• Journal of Environmental Science International
• /
• v.12 no.1
• /
• pp.77-80
• /
• 2003

The interaction of mastoparan B, a cationic tetradecapeptide amide isolated from the hornet Vespa basalis, with phospholipid bilayers was studied with synthetic mastoparan B and its analogue with Ala instead of hydrophobic 12th amino acid residue in mastoparan B. MP-B and its derivative, [12-Ala]MP-B were synthesized by the solid-phase peptide synthesis method. MP-B and its analogue, [12-Ala]MP-B adopted an unordered structure in buffer solution. In the presence of neutral and acidic liposomes, the peptides took an $\alpha$-helical structure. The two peptides interacted with neutral and acidic lipid bilayers. These results indicated that the hydrophobic face in the amphipathic $\alpha$-helix of MP-B critically affected the biological activity and helical content.

• Bulletin of the Korean Chemical Society
• /
• v.18 no.1
• /
• pp.50-56
• /
• 1997

The mode of action of mastoparan B, an antimicrobial cationic tetradecapeptide amide isolated from the hornet Vespa basalis, toward phospholipid bilayers was studied with synthetic mastoparan B and its analogs with individual Ala instead of hydrophobic amino acids (1-Ile, 3-Leu, 6-Leu, 7-Val, 9-Trp, 13-Val, 14-Leu) in mastoparan B. Mastoparan B and its analogs were synthesized by the solid-phase method. Circular dichroism spectra showed that mastoparan B and its analogs adopted an unordered structure in buffer solution. In the presence of neutral and acidic liposomes, most of the peptides took an α-helical structure. The calcein leakage experiment indicated that mastoparan B interacted strongly with neutral and acidic lipid bilayers than its analogs. Mastoparan B also showed a more or less highly antimicrobial activity and hemolytic activity for human erythrocytes than its analogs. These results indicate that the hydrophobic face in the amphipathic α-helix of mastoparan B critically affect biological activity and helical contents.