How C-Terminal Carboxyamidation Alters the Biological Activity of Peptides from the Venom of the Eumenine Solitary Wasp

Abstract

Inflammatory peptides display different types of post-transcriptional modifications, such as C-terminal amidation, that alter their biological activity. Here we describe the structural and molecular dynamics features of the mast cell degranulating peptide, eumenine mastoparan-AF (EMP-AF-NH(2)), found in the venom of the solitary wasp, and of its carboxyl-free C-terminal form (EMP-AF-COO(-)) characterized by a reduced activity. Circular dichroism indicates that both peptides switch from a random coil conformation in water to a helical structure in TFE and SDS micelles. NMR data, in 30% TFE, reveal that the two peptides fold into an alpha-helix spanning most of their length, while they differ in terms of molecular rigidity. To understand the origins of the conformational flexibility observed in the case of EMP-AF-COO(-), a 5 ns MD simulation was carried out for each peptide, in an explicit water/TFE environment. The results show that the two peptides differ in an H-bond between Leu14 NH(2) and the backbone carbonyl of Ile11. The loss of that H-bond in EMP-AF-COO(-) leads to a significant modification of its structural dynamics. In fact, as evidenced by essential dynamics analysis, while EMP-AF-NH(2) exists mainly as a rigid structure, EMP-AF-COO(-) presents two helical stretches that fluctuate in some sort of independent fashion. We conclude that the diverse biological activity of the two peptides is not simply due to the reduction of the net positive charge, as generally suggested, but also to a structural perturbation of the amphipathic alpha-helix that affects their ability to perturb the cell membrane.

Primary Structure and Antibacterial Activity of Chicken Bone Marrow-Derived β-Defensins

“…Elsewhere, the presence of pyroglutamic acid as the N-terminal residue, another kind of posttranslational modification, was found in the newly characterized AvBD7 sequence. Amidation and glutaminyl cyclization are commonly found in bioactive peptides and short polypeptides, such as hormones, neuropeptides, antimicrobial peptides, and venom toxins (6,21,24,27,29,32,41,42). The C-terminal amide has been shown to increase antibacterial potency by inducing ␣-helix formation or stabilizing ␣-helix (13,28,32).…”

Effect of C‐Terminal Amidation on Folding and Disulfide‐Pairing of α‐Conotoxin ImI

“…Such a modification has been shown to have a substantial impact on the biological activities of several of these peptides, presumably owing to the neutralization of negative charges conferred by the carboxylic acid at the C terminus. [22][23][24][25][26] In these cases, however, the C-terminal amide is not known to influence the conformation of the peptides. In earlier reports, post-translational modifications such as g-carboxylation of glutamate residues have been shown to be important for the folding in P-superfamily conotoxins.…”

Lipid-packing perturbation of model membranes by pH-responsive antimicrobial peptides

“…This C-terminal protection prevents proteolytic digestion of the peptide and has been shown to play an important role in the peptide structure. From the structural point of view, the C-terminus amidation was shown by NMR to provide an extra hydrogen bonding in Eumenine mastoparan-AF (EMP-AF-NH 2 ) compared with the non-amidated analog (Sforça et al 2004). Similar observations were carried out for Protonectarina-MP using hydrogen deuterium exchange and mass spectrometry (da Silva et al 2014).…”