Isolation and structural and pharmacological characterization of α-elapitoxin-Dpp2d, an amidated three finger toxin from black mamba venom.

We isolated a novel, atypical long-chain three-finger toxin (TFT), α-elapitoxin-Dpp2d (α-EPTX-Dpp2d), from black mamba (Dendroaspis polylepis polylepis) venom. Proteolytic digestion with trypsin and V8 protease, together with MS/MS de novo sequencing, indicated that the mature toxin has an amidated C-terminal arginine, a posttranslational modification rarely observed for snake TFTs. α-EPTX-Dpp2d was found to potently inhibit α7 neuronal nicotinic acetylcholine receptors (nAChR; IC₅₀, 58 ± 24 nM) and muscle-type nAChR (IC₅₀, 114 ± 37 nM) but did not affect α3β2 and α3β4 nAChR isoforms at 1 μM concentrations.

Semienzymatic cyclization of disulfide-rich peptides using Sortase A.

Disulfide-rich cyclic peptides have generated great interest in the development of peptide-based therapeutics due to their exceptional stability toward chemical, enzymatic, or thermal attack. In particular, they have been used as scaffolds onto which bioactive epitopes can be grafted to take advantage of the favorable biophysical properties of disulfide-rich cyclic peptides. To date, the most commonly used method for the head-to-tail cyclization of peptides has been native chemical ligation.

Chemical engineering and structural and pharmacological characterization of the α-scorpion toxin OD1.

Scorpion α-toxins are invaluable pharmacological tools for studying voltage-gated sodium channels, but few structure-function studies have been undertaken due to their challenging synthesis. To address this deficiency, we report a chemical engineering strategy based upon native chemical ligation. The chemical synthesis of α-toxin OD1 was achieved by chemical ligation of three unprotected peptide segments.

Efficient chemical synthesis of human complement protein C3a.

We report the total chemical synthesis of human C3a by one-pot native chemical ligation of three unprotected peptide segments, followed by efficient in vitro folding that yielded the anaphylatoxin C3a in high yield and excellent purity. Synthetic C3a was fully active and its crystal structure at 2.1 Å resolution showed 3 helices and a C-terminal turn motif.

Conopeptide ρ-TIA defines a new allosteric site on the extracellular surface of the α1B-adrenoceptor.

The G protein-coupled receptor (GPCR) superfamily is an important drug target that includes over 1000 membrane receptors that functionally couple extracellular stimuli to intracellular effectors. Despite the potential of extracellular surface (ECS) residues in GPCRs to interact with subtype-specific allosteric modulators, few ECS pharmacophores for class A receptors have been identified. Using the turkey β(1)-adrenergic receptor crystal structure, we modeled the α(1B)-adrenoceptor (α(1B)-AR) to help identify the allosteric site for ρ-conopeptide TIA, an inverse agonist at this receptor.

Emerging opportunities for allosteric modulation of G-protein coupled receptors.

Their ubiquitous nature, wide cellular distribution and versatile molecular recognition and signalling help make G-protein binding receptors (GPCRs) the most important class of membrane proteins in clinical medicine, accounting for ∼40% of all current therapeutics. A large percentage of current drugs target the endogenous ligand binding (orthosteric) site, which are structurally and evolutionarily conserved, particularly among members of the same GPCR subfamily. With the recent advances in GPCR X-ray crystallography, new opportunities for developing novel subtype selective drugs have emerged.

Conotoxin engineering: dual pharmacophoric noradrenaline transport inhibitor/integrin binding peptide with improved stability.

A dual-pharmacophoric peptide was engineered by grafting the integrin binding RGD motif between the C- and N-termini of a disulfide-rich noradrenaline transporter inhibiting χ-conotoxin resulting in a stable backbone cyclized peptide. The construct maintained two independent biological activities and showed increased plasma stability with no adverse effects observed following administration to rats, highlighting the potential value of pharmacophore grafting into constrained peptide scaffolds.

Atypical alpha-conotoxin LtIA from Conus litteratus targets a novel microsite of the alpha3beta2 nicotinic receptor.

Different nicotinic acetylcholine receptor (nAChR) subtypes are implicated in learning, pain sensation, and disease states, including Parkinson disease and nicotine addiction. alpha-Conotoxins are among the most selective nAChR ligands. Mechanistic insights into the structure, function, and receptor interaction of alpha-conotoxins may serve as a platform for development of new therapies.

Emerging structure-function relationships defining monoamine NSS transporter substrate and ligand affinity.

Monoamine transporters are a group of transmembrane neurotransmitter sodium symporter (NSS) transporters that play a crucial role in regulating biogenic monoamine concentrations at peripheral and central synapses. Given the key role played by serotonin, dopamine and noradrenaline in addictive and disease states, structure-function studies have been conducted to help guide the development of improved central nervous system therapeutics. Extensive pharmacological, immunological and biochemical studies, in conjunction with three-dimensional homology modeling, have been performed to structurally and functionally characterise the monoamine transporter substrate permeation pathway, substrate selectivity, and binding sites for ions, substrates and inhibitors at the molecular level.