The Effects of Structural Alterations in the Polyamine and Amino Acid Moieties of Philanthotoxins on Nicotinic Acetylcholine Receptor Inhibition in the Locust, Schistocerca gregaria

Alterations in the polyamine and amino acid (tyrosine) moieties of philanthotoxin‐343 (PhTX‐343) were investigated for their effects on the antagonism of nicotinic acetylcholine receptors (nAChRs) isolated from the locust (Schistocerca gregaria) mushroom body. Through whole‐cell patch‐clamp recordings, the philanthotoxin analogues in this study were shown to cause inhibition of the inward current when co‐applied with acetylcholine (ACh). PhTX‐343 (IC50 = 0.80 μM at −75 mV) antagonised locust nAChRs in a use‐dependent manner, suggesting that it acts as an open-channel blocker. The analogue in which both the secondary amine functionalities were replaced with methylene groups (i.e., PhTX‐12) was ~6‐fold more potent (IC50 (half‐maximal inhibitory con-centration) = 0.13 μM at −75 mV) than PhTX‐343. The analogue containing cyclohexylalanine as a substitute for the tyrosine moiety of PhTX‐343 (i.e., Cha‐PhTX‐343) was also more potent (IC50 = 0.44 μM at −75 mV). A combination of both alterations to PhTX‐343 generated the most potent analogue, i.e., Cha‐PhTX‐12 (IC50 = 1.71 nM at −75 mV). Modulation by PhTX‐343 and Cha‐PhTX‐343 fell into two distinct groups, indicating the presence of two pharmacologically distinct nAChR groups in the locust mushroom body. In the first group, all concentrations of PhTX‐343 and Cha‐PhTX‐343 inhibited responses to ACh. In the second group, application of PhTX‐343 or Cha‐PhTX‐343 at concentrations ≤100 nM caused potentiation, while concentrations ≥1 μM inhibited responses to ACh. Cha‐PhTX‐12 may have potential to be developed into insecticidal compounds with a novel mode of action

Controlled synthesis of O-glycopolypeptide polymers and their molecular recognition by lectins

The facile synthesis of high molecular weight water-soluble O-glycopolypeptide polymers by the ring-opening polymerization of their corresponding N-carboxyanhydride (NCA) in very high yield (overall yield > 70%) is reported. The per-acetylated-O-glycosylated lysine-NCA monomers, synthesized using stable glycosyl donors and a commercially available protected amino acid in very high yield, was polymerized using commercially available amine initiators. The synthesized water-soluble glycopolypeptides were found to be α-helical in aqueous solution. However, we were able to control the secondary conformation of the glycopolypeptides (α-helix vs nonhelical structures) by polymerizing racemic amino acid glyco NCAs. We have also investigated the binding of the glycopolypeptide poly(α-manno-O-lys) with the lectin Con-A using precipitation and hemagglutination assays as well as by isothermal titration calorimetry (ITC). The ITC results clearly show that the binding process is enthalpy driven for both α-helical and nonhelical structures, with negative entropic contribution. Binding stoichiometry for the glycopolypeptide poly(α-manno-O-lys) having a nonhelical structure was slightly higher as compared to the corresponding polypeptide which adopted an α-helical structure

Synthesis and Self-assembly of Amphiphilic Homoglycopolypeptide

The synthesis of the amphiphilic homoglycopolypeptide was carried out by a combination of NCA polymerization and click chemistry to yield a well-defined polypeptide having an amphiphilic carbohydrate on its side chain. The amphiphilicity of the carbohydrate was achieved by incorporation of an alkyl chain at the C-6 position of the carbohydrate thus also rendering the homoglycopolypeptide amphiphilic. The homoglycopolypeptide formed multimicellar aggregates in water above a critical concentration of 0.9 μM due to phase separation. The multimicellar aggregates were characterized by DLS, TEM, and AFM. It is proposed that hydrophobic interactions of the aliphatic chains at the 6-position of the sugar moieties drives the assembly of these rod-like homoglycopolypeptide into large spherical aggregates. These multimicellar aggregates encapsulate both hydrophilic as well as hydrophobic dye as was confirmed by confocal microscopy. Finally, amphiphilic random polypeptides containing 10% and 20% α-d-mannose in addition to glucose containing a hydrophobic alkyl chain at its 6 position were synthesized by our methodology, and these polymers were also found to assemble into spherical nanostructures. The spherical assemblies of amphiphilic random glycopolypeptides containing 10% and 20% mannose were found to be surface bioactive and were found to interact with the lectin Con-A