Structural and functional characterization of two genetically related meucin peptides highlights evolutionary divergence and convergence in antimicrobial peptides

Abstract

Both vertebrates and invertebrates employ alpha-helical antimicrobial peptides (AMPs) as an essential component of their innate immune system. However, evolutionary relation of these immune molecules remains unresolved. Venoms, as key weapons of venomous arthropods for prey and defense, receive increasing recognition as an emerging source of such peptides. From a cDNA library prepared from the venom gland of the scorpion Mesobuthus eupeus, clones encoding precursors of two new AMPs, named meucin-13 (IFGAIAGLLKNIF-NH(2)) and meucin-18 (FFGHLFKLATKIIPSLFQ), have been isolated. The precursor of meucins consists of a signal peptide, a mature peptide, and an acidic propeptide, in which dibasic residues as the typical processing signal are located between the mature and propeptide. Meucin-13 is an ortholog of several previously described AMPs from scorpion venom and has also detectable sequence similarity to temporins, a large family of AMPs from frog skin, whereas meucin-18 displays some similarity to AMPs from diverse origin including arthropod venoms, fish mast cells, and frog skins. These two meucin peptides form alpha-helical structure in the presence of 50% trifluoroethanol (TFE), a membrane-mimicking environment, as identified by circular dichroism (CD) spectroscopy. This finding is further verified by their NMR structures that show a typical alpha-helical amphipathic design, a structural prerequisite for cytolytic activity. Meucins exhibit extensive cytolytic effects on both prokaryotic and eukaryotic cells (gram(+) and gram(-) bacteria, fungi, yeasts, rabbit erythrocytes, and rat dorsal root ganglion cells) at micromolar concentrations. It is remarkable that muecin-18 was 2- to >14-fold more potent than meucin-13 against nearly all the cells tested. Structural differences in hydrophilic/hydrophobic balance and cationic amino acid location between two meucins could account for their differential potency. Despite these differences, commonalities at precursor organization, three-dimensional structure, and biological function suggests that meucins are two evolutionarily related AMPs and likely originated from a common ancestor by gene duplication. Our work presented here also provides new insights into an evolutionary link among AMPs from invertebrates and vertebrates and clues for evolutionary convergence between AMPs and virus fusion domains.

Dermatophytic defensin with antiinfective potential

“…On the contrary, meucin-18 caused an immediate fluorescence increase upon exposure of the peptide. These observations are in agreement with features of these two control molecules previously reported (20,21). At a concentration up to 20× C L , micasin only led to weak dye uptake after 6 min.…”

“…Analysis of the structures with PROMOTIF (17) identified an α-helix between residues 7-18 and a β-sheet consisting of two antiparallel strands between residues 22-26 and 32-36. A short sequence of only three residues (20)(21)(22) links the helix to strand I. The loop connecting the two β-strands (residues 27-31) is highly flexible, with no dominant turn type defined.…”

“…The ability of micasin to permeabilize the bacterial membrane was also assessed with the fluorescent nucleic acid-binding dye propidium iodide. Vancomycin and meucin-18 (a pore-forming peptide from scorpion venom) were used as negative and positive control (20,21). No fluorescence increase was observed over 10 min after B. megaterium were exposed to micasin and vancomycin at 5× C L or 10× C L (Fig.…”

Exploring the global animal biodiversity in the search for new drugs – Spiders, scorpions, horseshoe crabs, sea spiders, centipedes, and millipedes

“…Among these are not only ion channel modulators [137,138] but also a number of membrane-active antimicrobial peptides called meucins. These compounds exhibited extensive cytolytic effects on, among others, Gram-positive and Gramnegative bacteria at micromolar concentrations [139]. Like many other antimicrobial peptides, meucins are believed to represent ancient biological weapons against bacteria that selectively target the bacterial cell surface [140].…”

Insights into Antimicrobial Peptides from Spiders and Scorpions

“…All of the three peptides possess stronger activities against Gram-positive bacteria than Gram-negative bacteria. Meucin-13 and meucin-18 were isolated from a cDNA library of the venom gland of the scorpion Mesobuthus eupeus [75]. They showed sequence similarity to amphibian 13-residue temporins and 18-residue brevinins, respectively.…”