1.C.16 The Magainin (Magainin) Family

Many organisms synthesize proteins (or peptides) which are degraded to relatively small hydrophobic or amphipathic, bioactive peptides. These peptides exhibit antibiotic, fungicidal, virucidal, hemolytic and/or tumoricidal activities by interacting with membranes and forming transmembrane channels that allow the free flow of electrolytes, metabolites and water across the phospholipid bilayers. Most of these peptides appear to function in biological warfare. There are many designations given to these bioactive peptides. They include the magainins, cecropins, melittins, defensins, bacteriocidins, etc. The proteins in each family within this functional superfamily are homologous, but they exhibit little or no significant sequence similarity with members of the other families. Thus, each family may have evolved independently. However, certain common structural features observed between members of distinct families suggest that at least some of these families share a common ancestry. Pore size and subunit stoichiometry of the pore have been determined (Watanabe and Kawano 2016).

Magainins are a group of amphipathic peptides (21-26 amino acyl residues in length) from the skin and intestines of frogs. They form right-handed α-helical structures in membranes and serve bactericidal, fungicidal and virucidal functions. They thereby provide defense against infectious agents. As revealed by their synergistic behavior, they can form homo- or heterooligomeric transmembrane channels. One distant member of the magainin family, 'peptide between glycine and leucine amide' (PGLa) (GMASKAGAIAGKIAKVALKAL-NH2) is not hemolytic but acts on Gram-negative and Gram-positive bacteria as well as fungi and protozoa. PGLa is positively charged, adopts a random coil configuration in water and folds into an amphipathic α-helix when it inserts into the membrane, parallel to the membrane surface at low peptide concentrations but perpendicular to the membrane surface at higher concentrations when it forms a peptide/lipid pore. Pore formation depends on anionic phospholipids. PGLa and magainin2 amide exhibit synergism when forming transmembrane pores. The synergism of magainin 2 amide and PGLa in forming transmembrane pores results from formation of a heterodimer composed of parallel helices with strong membrane permeabilizing activity (Hara et al., 2001; Nishida et al., 2007). While magainin 1 is anion-selective, magainin 2 is cation-selective (Kourie and Shorthouse, 2000).

The antimicrobial peptides magainin 2 and the PGLa (PYLa) precursor (TC# 1.C.16.1.5) show marked synergism. These two peptides form a heterodimer composed of parallel helices with strong membrane permeabilizing activity (Hara et al., 2001). The synergy observed for the two peptides is due to the formation of a parallel heterodimer (Nishida et al., 2007). Magainin disruption of cells is concentration-dependent and a highly stochastic process (Han et al., 2009). Based on NMR and other analyses, Han et al., 2009 concluded that magainin-induced pores in lipid vesicles have a mean diameter of approximately 80 A.

Model membranes have been used to elucidate how peptides permeabilize membranes. The interaction of F5W-magainin 2 (GIGKWLHSAKKFGKAFVGEIMNS), an equipotent analogue of magainin 2 isolated from the African clawed frog Xenopus laevis was studied with unfixed Bacillus megaterium and Chinese hamster ovary (CHO)-K1 cells by Imura et al. (2008). The influx of fluorescent markers of various sizes into the cytosol revealed that magainin 2 permeabilizes bacterial and mammalian membranes in significantly different ways. The peptide formed pores with a diameter of approximately 2.8 nm (<6.6 nm) in B. megaterium. In contrast, the peptide perturbed the membrane of CHO-K1 cells, permitting the entry of large molecules (diameter, >23 nm) into the cytosol. 

Magainin 2 and PGLa bind preferentially to negatively charged membranes and cause their disruption by pore formation. All-atom molecular dynamics simulations starting from tetrameric transmembrane helical bundles of these two peptides, as well as their stoichiometric mixture produced pore structures (Pino-Angeles et al. 2016). The peptides remained mostly helical and adopted tilted orientations. The calculated tilt angles for PGLa were in agreement with solid state NMR experiments. The antiparallel dimer structure in the magainin 2 simulations resembled previously determined NMR and crystal structures. A larger and more ordered pore was seen in the 1:1 heterotetramer with an antiparallel helix arrangement. 

Magainin 2 (MAG2) and PGLa, in the skin of the African frog Xenopus laevis, act by permeabilizing bacterial membranes and exhibit synergism. The amphiphilic MAG2 helix lies flat on the membrane surface in some lipid membranes, but with a tilt angle close to 90 degrees (Strandberg et al. 2016).  In the presence of an equimolar amount of PGLa, MAG2 becomes tilted at an angle of 120 degrees, and its azimuthal rotation angle changes. Since this interaction was found to occur in a concentration range where the peptides per se do not interact with their own type, it seems that MAG2 forms a stable heterodimer with PGLa. Given that PGLa molecules in the complex are known to be flipped into a fully upright orientation, with a helix tilt close to 180 degrees , they must make up the actual transmembrane pore (Strandberg et al. 2016). Possibly the two negative charges on the C-terminus of the obliquely tilted MAG2 peptides neutralize some of the cationic groups on the upright PGLa helices. This would stabilize the assembly of PGLa into a toroidal pore with an overall reduced charge density, which could explain the mechanism of synergy.

The generalized transport reaction catalyzed by channel-forming amphipathic peptides is:

Small solutes, electrolytes and water (in) small solutes, electrolytes and water (out)



This family belongs to the .

 

References:

Bechinger, B. (1997). Structure and functions of channel-forming peptides: magainins, cecropins, melittin and alamethicin. J. Membr. Biol. 156: 197-211.

Bechinger, B., M. Zasloff, and S.J. Opella. (1993). Structure and orientation of the antibiotic peptide magainin in membrane by solid-state nuclear magnetic resonance spectroscopy. Prot. Sci. 2: 2077-2084.

Han, M., Y. Mei, H. Khant, and S.J. Ludtke. (2009). Characterization of antibiotic peptide pores using cryo-EM and comparison to neutron scattering. Biophys. J. 97: 164-172.

Hara, T., Y. Mitani, K. Tanaka, N. Uematsu, A. Takakura, T. Tachi, H. Kodama, M. Kondo, H. Mori, A. Otaka, F. Nobutaka, and K. Matsuzaki. (2001). Heterodimer formation between the antimicrobial peptides magainin 2 and PGLa in lipid bilayers: a cross-linking study. Biochemistry. 40: 12395-12399.

Imura, Y., N. Choda, and K. Matsuzaki. (2008). Magainin 2 in action: distinct modes of membrane permeabilization in living bacterial and mammalian cells. Biophys. J. 95: 5757-5765.

Kourie, J.I. and A.A. Shorthouse. (2000). Properties of cytotoxic peptide-formed ion channels. Am. J. Physiol. Cell Physiol. 278: C1063-C1087.

Matsuzaki, K. (1998). Magainins as paradigm for the mode of action of pore forming polypeptides. Biochim. Biophys. Acta 1376: 391-400.

Matsuzaki, K. (1999). Why and how are peptide-lipid interactions utilized for self-defense? Magainins and tachyplesins as archetypes. Biochim. Biophys. Acta 1462: 1-10.

Münster, C., A. Spaar, B. Bechinger, and T. Salditt. (2002). Magainin 2 in phospholipid bilayers: peptide orientation and lipid chain ordering studied by x-ray diffraction. Biochim. Biophys. Acta 1562: 37-44.

Nishida, M., Y. Imura, M. Yamamoto, S. Kobayashi, Y. Yano, and K. Matsuzaki. (2007). Interaction of a magainin-PGLa hybrid peptide with membranes: insight into the mechanism of synergism. Biochemistry. 46: 14284-14290.

Parvez, F., J.M. Alam, H. Dohra, and M. Yamazaki. (2018). Elementary processes of antimicrobial peptide PGLa-induced pore formation in lipid bilayers. Biochim. Biophys. Acta. Biomembr 1860: 2262-2271.

Pino-Angeles, A., J.M. Leveritt, 3rd, and T. Lazaridis. (2016). Pore Structure and Synergy in Antimicrobial Peptides of the Magainin Family. PLoS Comput Biol 12: e1004570.

Strandberg, E., D. Horn, S. Reißer, J. Zerweck, P. Wadhwani, and A.S. Ulrich. (2016). (2)H-NMR and MD Simulations Reveal Membrane-Bound Conformation of Magainin 2 and Its Synergy with PGLa. Biophys. J. 111: 2149-2161.

Tanphaichitr, N., N. Srakaew, R. Alonzi, W. Kiattiburut, K. Kongmanas, R. Zhi, W. Li, M. Baker, G. Wang, and D. Hickling. (2016). Potential Use of Antimicrobial Peptides as Vaginal Spermicides/Microbicides. Pharmaceuticals (Basel) 9:.

Ulmschneider, J.P. (2017). Charged Antimicrobial Peptides Can Translocate across Membranes without Forming Channel-like Pores. Biophys. J. 113: 73-81.

Watanabe, H. and R. Kawano. (2016). Channel Current Analysis for Pore-forming Properties of an Antimicrobial Peptide, Magainin 1, Using the Droplet Contact Method. Anal Sci 32: 57-60.

Wieprecht, T., O. Apostolov, M. Beyermann, and J. Seelig. (2000). Membrane binding and pore formation of the antibacterial peptide PGLa: thermodynamic and mechanistic aspects. Biochemistry. 39: 442-452.

Examples:

TC#NameOrganismal TypeExample
1.C.16.1.1

Magainin precursor of 333 aas and 1 TMS.  3-d structural determinations and simulations show the oligomeric states, transmembrane helices and tilt angles in the various states of the mature Maganin (Pino-Angeles et al. 2016).  Forms stable heterooligomers with PglA (TC# 1.C.16.1.5) at lower concentrations of the two peptides than allows each one alone to form pores in which PglA, rather than magainin 2 forms the pore (Strandberg et al. 2016).

Frogs

Magainin precursor of Xenopus laevis

 
1.C.16.1.2Preprocaerulein Frogs Preprocaerulein type I of Xenopus laevis
 
1.C.16.1.3Xenopsin precursor Frogs Xenopsin precursor of Xenopus laevis
 
1.C.16.1.4Prolevitide precursor Frogs Prolevitide precursor of Xenopus laevis
 
1.C.16.1.5

PylA/PglA (peptide glycine-leucine-amide) precursor of 64 aas and 1 TMS.  3-d structures and simulations have revealed the overall structure, helix orientations, and tilt angles in the homo- and hetero-multimeric pores (Pino-Angeles et al. 2016).  The pore forms stable heterooligomers with magainin 2 (TC# 1.C.16.1.1) in which PglA, rather than magainin 2, forms the pore (Strandberg et al. 2016).  This occurs at lower concentrations of the two peptides than is required for each peptide to form homomeric pores.  Ulmschneider 2017 suggested that cationic antimicrobial peptides (AMPs) such as PGLa translocate across hydrophobic lipid bilayers without formation of peptide-lined channels, explaining why they induce membrane leakage and antimicrobial activity. PGLa spontaneously translocates across the membrane individually on a timescale of tens of microseconds, without forming pores. Instead, short-lived water bridges, with two or three peptides connecting at their termini, may allow both ion translocation and lipid flip-flop via a brushlike mechanism usually involving the C terminus of one peptide (Ulmschneider 2017). Another study suggested that PGLa translocates across the bilayer before membrane permeation (Parvez et al. 2018).

Frogs

PylA/PglA precursor of Xenopus laevis

 
1.C.16.1.6

Toxic magainin peptide, Magainin-R-2 of 23 aas. Magainins are membrane lytic agents.  From the parent protein (1.C.16.1.1), many antimicrobial peptides that inhibit the growth of numerous species of bacteria and fungi and induce osmotic lysis of protozoa can be derived (Tanphaichitr et al. 2016).

Magainin-R-2 of Xenopus laevis (African clawed frog)

 
Examples:

TC#NameOrganismal TypeExample
1.C.16.2.1Hypothetical Protein (99aas)

Alveolata

Hypothetical protein of Toxoplasma gondii (B6K9W1)

 
1.C.16.2.2

Uncharacterized protein of 99 aas and 1 TMS.

UP of Hammondia hammondi

 
1.C.16.2.3

Uncharacterized protein of 93 aas and 1 TMS.

UP of Neospora caninum