Cupricyclins, Novel Redox-Active Metallopeptides Based on Conotoxins Scaffold

Abstract

Highly stable natural scaffolds which tolerate multiple amino acid substitutions represent the ideal starting point for the application of rational redesign strategies to develop new catalysts of potential biomedical and biotechnological interest. The knottins family of disulphide-constrained peptides display the desired characteristics, being highly stable and characterized by hypervariability of the inter-cysteine loops. The potential of knottins as scaffolds for the design of novel copper-based biocatalysts has been tested by engineering a metal binding site on two different variants of an ω-conotoxin, a neurotoxic peptide belonging to the knottins family. The binding site has been designed by computational modelling and the redesigned peptides have been synthesized and characterized by optical, fluorescence, electron spin resonance and nuclear magnetic resonance spectroscopy. The novel peptides, named Cupricyclin-1 and -2, bind one Cu2+ ion per molecule with nanomolar affinity. Cupricyclins display redox activity and catalyze the dismutation of superoxide anions with an activity comparable to that of non-peptidic superoxide dismutase mimics. We thus propose knottins as a novel scaffold for the design of catalytically-active mini metalloproteins.

A Disulfide Stabilized β-Sandwich Defines the Structure of a New Cysteine Framework M-Superfamily Conotoxin

“…Another aspect of the structure and stability of these molecules that has emerged has been their use as structural scaffolds upon which peptides with new functionality have been designed. [4][5][6] To date, ∼2100 conotoxins, categorized into 26 structurally distinct cysteine frameworks (arrangement of cysteines in the primary structure), 7 have been identified either as peptides in the venom or as amino acid sequences that have been derived from transcriptome data. 8 Tertiary structures are known for 65 wild-type conotoxins, and they are representative of 10 of the 26 known cysteine frameworks.…”

“…In general, bioactive peptides from venom of animals are cysteine rich, and the multiple disulfide bonds reinforce the stability. Another aspect of the structure and stability of these molecules that has emerged has been their use as structural scaffolds upon which peptides with new functionality have been designed. - …”

Snails In Silico: A Review of Computational Studies on the Conopeptides

“…They identified α-BuIA as strongly binding to LPAR6, making it a good candidate for investigation and refinement as a possible anti-cancer drug. Gao et al [167] performed a homology search of Conus betulinus venoms to identify six sequences similar to α-ImI, of which two were demonstrated to have desirable insecticidal properties, while Barba et al [168] performed a sequence scan followed by MD simulations as the starting point for the design of an ω-GVIA mutant that strongly binds copper atoms to be used for environmental applications. Using conantokin-G as a starting sequence, Reyes-Guzman et al [169] employed docking studies to evaluate mutants and guide a search that resulted in two peptides (EAR16 and EAR18) that are capable of reversibly blocking the GluN2B NMDA receptor, which is implicated in neuronal function.…”

Constant-pH MD Simulations Portray the Protonation and Structural Behavior of Four Decapeptides Designed to Coordinate Cu2+

“…Short peptides containing histidines may bind Cu 2+ very effectively, − having been used as model systems to study the role of Cu 2+ in the misfolding of proteins related to neurodegenerative disorders − and to mimic the structural and catalytic features of active sites in metalloenzymes. ,− In order to be good metalloprotein mimetics, these peptides should be capable of coordinating Cu 2+ exclusively through their side chains and forming single complexation species with appropriated coordination pockets at physiological pH. − However, most of these peptides at neutral and slightly basic pH form coordination species involving the main chain amide nitrogens as well as the imidazole pyrrole nitrogens, the latter acting as primary anchoring sites for Cu 2+ and promoting the deprotonation of the amides and their subsequent binding to the metal ions. − Hence, multihistidine peptides capable of binding Cu 2+ exclusively through the imidazole rings at neutral pH are still scarce. ,, …”