The primary target of Magainin 2 is the lipid matrix of cell membranes, specifically anionic phospholipids such as phosphatidylglycerol (PG), phosphatidylserine (PS), and phosphatidic acid (PA). The peptide does not require chiral proteinaceous receptors, as all-D enantiomers are fully active. [2]
The interaction with acidic lipopolysaccharides (LPS) on the outer surface of Gram-negative bacteria is also a target. [2]
The peptide dissipates the electric potential across energy-transducing membranes, uncoupling respiration from energy-requiring processes. [2]

Magainin 2 causes morphological alterations during early apoptosis in E. coli and demonstrates bactericidal effects. coli. RecA's expression as a caspase-like protein is influenced by Magainin 2, which also causes the expression of bacterial proteins that have an affinity for caspase substrates [1]. By permeabilizing cell membranes, magainin 2 kills bacteria without posing a serious threat to mammalian cells. It is believed that the lipid matrix of the membrane is the primary target of the peptide. When natural Paramecium caudatum was exposed to 10 μg/mL magainin 2, it caused osmotic swelling and eventual cell rupture in the pond water. This suggests that the peptide may interfere with the membrane function that maintains osmotic balance [2]. Magainin 2 permeabilizes cell membranes in mammals and bacteria in distinctly different ways. In Bacillus megaterium, the peptide produces holes with a diameter of around 2.8 nm (less than 6.6 nm) and translocates into the cytoplasm. The CHO-K1 cell membrane, on the other hand, is severely disrupted by this peptide, allowing big molecules (greater than 23 nm) to enter the cytoplasm along with membrane budding and lipid flipping, which mostly causes the molecules to accumulate in the nucleus and mitochondria [3].

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Magainin 2 exhibited antibiotic activity against numerous Gram-negative and Gram-positive bacteria. Minimal inhibitory concentrations (MICs) were: Klebsiella pneumoniae 10 μg/ml, Pseudomonas putida 10 μg/ml, Staphylococcus epidermidis 10 μg/ml, Citrobacter freundii 30 μg/ml, Enterobacter cloacae 50 μg/ml, Escherichia coli 50 μg/ml, Staphylococcus aureus 50 μg/ml, Candida albicans 80 μg/ml, Pseudomonas aeruginosa 100 μg/ml, Serratia marcescens 100 μg/ml, while Proteus mirabilis and Streptococcus fecalis showed resistance at >100 μg/ml. [1]
Magainin 2 was bactericidal against E. coli D31: at 10 μg/ml, it irreversibly reduced viability. [1]
Magainin 2 at 10 μg/ml caused osmotic swelling and subsequent bursting of Paramecium caudatum within minutes, with preserved swimming behavior (ciliary function) until swelling. Similar effects were observed on Amoeba proteus and Euglena gracilis. [1]
Magainin 2 was not hemolytic against human erythrocytes up to at least 150 μg/ml in phosphate-buffered saline, whereas melittin (a 26-amino acid amphiphilic peptide from bee venom) caused hemolysis at much lower concentrations. [1]
Magainin 2 binds preferentially to and permeabilizes bilayers containing acidic phospholipids (PG, PS, PA) via electrostatic interactions. Its affinity for zwitterionic phosphatidylcholine (PC) is extremely weak. Incorporation of cholesterol into PS bilayers inhibits membrane permeabilization activity. [2]
Magainin 2 forms pores or structural defects of approximately 1 nm diameter in liposomes and mitochondrial inner membranes, allowing passage of molecules up to molecular weight 623 Da (calcein) but not trypsin (24 kDa). The pore lifetime strongly depends on the peptide net charge. [2]
The membrane permeabilization by Magainin 2 depends on the 3rd to 6th power of the peptide concentration, indicating involvement of several peptide molecules (average of five helices per pore). The pore formation is transient and coupled to peptide translocation into the inner leaflet, which deactivates the pore. [2]
Magainin 2 induces a measurable extent of dye release from egg PG large unilamellar vesicles (LUVs) under conditions where several hundred peptide molecules are bound per vesicle, suggesting the active fraction is ~1%. [2]
The charge distribution of wild-type Magainin 2 (net +4) is optimized to exhibit maximal lytic activity by achieving strong binding and moderate pore stability. Increasing net positive charge enhances binding to negatively charged membranes but destabilizes the pore; decreasing charge reduces binding efficiency. [2]
The angle subtended by the positively charged helix face modulates pore formation: increasing the angle from 80° to 180° decreased pore formation activity by a factor of 5. [2]

The standard antibacterial assay used E. coli D31. Bacteria were grown in LB broth to an OD600 of 0.8 (representing 10^9 colony-forming units/ml), and 10^6 bacteria were added to 8 ml of 0.7% agarose in LB broth poured over a 150-mm Petri dish containing 50 ml of 1.5% agarose in LB broth. Antibacterial activity was assayed by suppression of bacterial growth dependent on application of fractions to the top agar surface. Other organisms were assayed in this manner or in liquid culture. For liquid culture assays, fractions were added to 100 μl of a suspension of organisms diluted from midlogarithmic-phase liquid culture to 10^5 cells per ml in standard TSB broth (pH 7.5). After incubation at 37°C for 4 hr, OD600 was measured. [1]
For hemolytic assay, either Magainin 2 or melittin was added to 100 μl of a 10% (vol/vol) suspension of human erythrocytes in phosphate-buffered saline. Samples were incubated at 37°C for 10 min, centrifuged at 10,000 × g for 10 min to remove cells and debris, and hemolysis was determined by measurement of OD350 of the supernatant. Addition of 0.1% Triton X-100 defined 100% hemolysis. [1]
For protozoan assay, Paramecium caudatum were exposed to Magainin 2 at 10 μg/ml in pond water or 1% TSB in distilled water; swelling of contractile vacuoles and subsequent cell bursting were observed. [1]
For fluorescence quenching experiments to determine helix orientation, three analog peptides of Magainin 2 each having a Trp residue substituted for Phe at position 5, 12, or 16 were synthesized. These derivatives were confirmed to be fully active compared to the parent peptide. Fluorescence quenching using doxylphosphatidylcholines revealed that each Trp residue is located approximately 1 nm from the bilayer center, suggesting the helix lies in the head group region. [2]
For calcein release assays, the self-quenching property of calcein (MW 623) was used to estimate pore lifetime and membrane permeabilization. Magainin 2 was added to liposomes, and dye release kinetics were monitored. The observed dye efflux deviated downward from first-order kinetics and ceased within approximately 10 min, indicating pore deactivation. [2]
For translocation detection, F12W Magainin 2 was incubated with labeled liposomes, and untranslocated peptide remaining on the outer leaflet was removed by either trypsin digestion or extraction with a large excess of unlabeled liposomes. Translocation was detected using fluorescence resonance energy transfer between the Trp residue of the peptide and dansyl-PE incorporated in the membrane. [2]

Magainin 2 is nonhemolytic at its effective antimicrobial concentrations. In hemolytic assays using human erythrocytes, Magainin 2 caused no hemolysis up to at least 150 μg/ml in phosphate-buffered saline, whereas melittin (a hemolytic amphiphilic peptide) was hemolytic at much lower concentrations. [1]
Magainin 2 exhibits no significant toxicity against mammalian cells. The selective toxicity is at least partly explained by preferential interactions with anionic phospholipids abundant in bacterial membranes, while mammalian cell membranes are composed of zwitterionic phospholipids and contain considerable cholesterol, which inhibits magainin activity. [2]

[1]. Magainins, a class of antimicrobial peptides from Xenopus skin: isolation, characterization of two active forms, and partial cDNA sequence of a precursor. Proc Natl Acad Sci U S A. 1987 Aug;84(15):5449-53.

[2]. Magainins as paradigm for the mode of action of pore forming polypeptides. Biochim Biophys Acta. 1998 Nov 10;1376(3):391-400.

According to reports, the African clawed frog contains magainin 2 peptide, and relevant data is available for reference.

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The name "magainin" is derived from the Hebrew word "magain" meaning "shield", reflecting its possible function as an antimicrobial shield. [1]
The primary sequence of Magainin 2 (23 amino acids) is: G I G K F L H S A K K F G K A F V G E I M N S (residues that differ from magainin 1 are underlined; magainin 1 differs by two substitutions). [1]
A computer search revealed no significant homology of Magainin 2 to any prokaryotic or eukaryotic protein in the GenBank file. [1]
Partial cDNA sequence of the precursor showed that Magainin 2 is encoded within a larger protein containing three magainin sequences, each bracketed by putative proteolytic cleavage sites (an arginine at the amino terminus and a Lys-Arg dipeptide at the carboxyl terminus) and common leader and trailer sequences of 6 and 7 amino acids respectively. [1]
Magainin 2 adopts an α-helical conformation upon binding to acidic phospholipid bilayers, with helicity ranging from 60-90% depending on experimental conditions. In aqueous neutral pH solutions, it has little or no well-defined secondary structure. [2]
Solid-state NMR and fluorescence quenching studies indicate that the Magainin 2 helix lies parallel to the bilayer surface (surface orientation) at low peptide-to-lipid ratios, but starts to insert perpendicular to the membrane surface above a peptide-to-lipid ratio of 1:30. [2]
Magainin 2 forms oligomers (dimer or trimer) above a bound peptide-to-lipid ratio of 0.02, based on sigmoidal binding isotherms of Trp-substituted analogs to PG membranes. At lower pH or high salt concentration (0.5 M NaCl), fibrils of 13 nm diameter are observed. [2]
The pore structure of Magainin 2 is of a torus type where lipids bend back on themselves (like the inside of a torus), as revealed by neutron diffraction, while the alamethicin pore is a barrel-stave type. The pore lining is composed of both the polar face of the amphiphilic helix and the polar heads of lipids, allowing lateral diffusion of lipid molecules between leaflets. [2]
The application of an inside-negative transmembrane potential electrophoretically facilitates translocation of the cationic Magainin 2. [2]
Magainin 2 triggers coupled transbilayer transport of ions, lipids, and the peptide itself (flip-flop). The lipid flip-flop rate is independent of the lipid head group structure, and inward and outward rates of lipid movement are identical, while the peptide translocates almost unidirectionally into the inner leaflet. [2]

C114H180N30O29S

2466.8976

2465.33

108433-95-0

16130189

White to off-white solid powder

4.913

33

36

86

174

5180

21

CC[C@H](C)[C@@H](C(=O)NCC(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC2=CN=CN2)C(=O)N[C@@H](CO)C(=O)N[C@@H](C)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC3=CC=CC=C3)C(=O)NCC(=O)N[C@@H](CCCCN)C(=O)N[C@@H](C)C(=O)N[C@@H](CC4=CC=CC=C4)C(=O)N[C@@H](C(C)C)C(=O)NCC(=O)N[C@@H](CCC(=O)O)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](CC(=O)N)C(=O)N[C@@H](CO)C(=O)O)NC(=O)CN

MGIUUAHJVPPFEV-ABXDCCGRSA-N

InChI=1S/C114H180N30O29S/c1-12-65(7)94(142-88(148)55-119)112(170)124-59-90(150)129-74(38-24-28-45-116)100(158)137-81(51-70-33-19-15-20-34-70)106(164)135-79(49-63(3)4)105(163)138-83(53-72-56-121-62-125-72)107(165)140-85(60-145)110(168)127-67(9)96(154)131-75(39-25-29-46-117)101(159)132-76(40-26-30-47-118)102(160)136-80(50-69-31-17-14-18-32-69)98(156)122-57-89(149)128-73(37-23-27-44-115)99(157)126-68(10)97(155)134-82(52-71-35-21-16-22-36-71)109(167)143-93(64(5)6)111(169)123-58-91(151)130-77(41-42-92(152)153)104(162)144-95(66(8)13-2)113(171)133-78(43-48-174-11)103(161)139-84(54-87(120)147)108(166)141-86(61-146)114(172)173/h14-22,31-36,56,62-68,73-86,93-95,145-146H,12-13,23-30,37-55,57-61,115-119H2,1-11H3,(H2,120,147)(H,121,125)(H,122,156)(H,123,169)(H,124,170)(H,126,157)(H,127,168)(H,128,149)(H,129,150)(H,130,151)(H,131,154)(H,132,159)(H,133,171)(H,134,155)(H,135,164)(H,136,160)(H,137,158)(H,138,163)(H,139,161)(H,140,165)(H,141,166)(H,142,148)(H,143,167)(H,144,162)(H,152,153)(H,172,173)/t65-,66-,67-,68-,73-,74-,75-,76-,77-,78-,79-,80-,81-,82-,83-,84-,85-,86-,93-,94-,95-/m0/s1

(4S)-4-[[2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-6-amino-2-[[2-[[(2S)-2-[[(2S)-6-amino-2-[[(2S)-6-amino-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-6-amino-2-[[2-[[(2S,3S)-2-[(2-aminoacetyl)amino]-3-methylpentanoyl]amino]acetyl]amino]hexanoyl]amino]-3-phenylpropanoyl]amino]-4-methylpentanoyl]amino]-3-(1H-imidazol-5-yl)propanoyl]amino]-3-hydroxypropanoyl]amino]propanoyl]amino]hexanoyl]amino]hexanoyl]amino]-3-phenylpropanoyl]amino]acetyl]amino]hexanoyl]amino]propanoyl]amino]-3-phenylpropanoyl]amino]-3-methylbutanoyl]amino]acetyl]amino]-5-[[(2S,3S)-1-[[(2S)-1-[[(2S)-4-amino-1-[[(1S)-1-carboxy-2-hydroxyethyl]amino]-1,4-dioxobutan-2-yl]amino]-4-methylsulfanyl-1-oxobutan-2-yl]amino]-3-methyl-1-oxopentan-2-yl]amino]-5-oxopentanoic acid

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture and light.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

H2O : ~50 mg/mL (~20.27 mM)

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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 (Please use freshly prepared in vivo formulations for optimal results.)