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Conformational Analysis of the Host-Defense Peptides Pseudhymenochirin-1Pb and -2Pa and Design of Analogues with Insulin-Releasing Activities and Reduced Toxicities

Manzo, Giorgia, Scorciapino, Mariano Andrea, Srinivasan, Dinesh, Attoub, Samir, Mangoni, Maria Luisa, Rinaldi, Andrea C., Casu, Mariano, Flatt, Peter R., Conlon, J. Michael
Journal of natural products 2015 v.78 no.12 pp. 3041-3048
amino acids, amphibians, antimicrobial properties, cytotoxicity, hemolysis, humans, inhibitory concentration 50, insulin, islets of Langerhans, molecular conformation, neoplasm cells, noninsulin-dependent diabetes mellitus, peptides, plasma membrane, solvents, therapeutics
Pseudhymenochirin-1Pb (Ps-1Pb; IKIPSFFRNILKKVGKEAVSLIAGALKQS) and pseudhymenochirin-2Pa (Ps-2Pa; GIFPIFAKLLGKVIKVASSLISKGRTE) are amphibian peptides with broad spectrum antimicrobial activities and cytotoxicity against mammalian cells. In the membrane-mimetic solvent 50% (v/v) trifluoroethanol–H₂O, both peptides adopt a well-defined α-helical conformation that extends over almost all the sequence and incorporates a flexible bend. Both peptides significantly (p < 0.05) stimulate the rate of release of insulin from BRIN-BD11 clonal β-cells at concentrations ≥ 0.1 nM but produce loss of integrity of the plasma membrane at concentrations ≥ 1 μM. Increasing cationicity by the substitution Glu¹⁷ → l-Lys in Ps-1Pb and Glu²⁷ → l-Lys in Ps-2Pa generates analogues with increased cytotoxicity and reduced insulin-releasing potency. In contrast, the analogues [R8r]Ps-1Pb and [K8k,K19k]Ps-2Pa, incorporating d-amino acid residues to destabilize the α-helical domains, retain potent insulin-releasing activity but are nontoxic to BRIN-BD11 cells at concentrations of 3 μM. [R8r]Ps-1Pb produces a significant increase in insulin release rate at 0.3 nM and [K8k,K19k]Ps-2Pa at 0.01 nM. Both analogues show low hemolytic activity (IC₅₀ > 100 μM) but retain broad-spectrum antimicrobial activity and remain cytotoxic to a range of human tumor cell lines, albeit with lower potency than the naturally occurring peptides. These analogues show potential for development into agents for type 2 diabetes therapy.