ABSTRACT
During the long-term evolution of animal toxins acting on potassium channels, the acidic residues can orientate the toxin binding interfaces by adjusting the molecular polarity. Based on the evolutionary function of toxin acidic residues, de novo peptide drugs with distinct binding interfaces were designed for the immunotherapeutic target, the Kv1.3 channel. Using a natural basic toxin, BmKTX, as a template, which contains 2 acidic residues (Asp19 and Asp33), we engineered two new peptides BmKTX-19 with 1 acidic residue (Asp33), and BmKTX-196 with 2 acidic residues (Asp6 and Asp33) through only adjusting acidic residue distribution for reorientation of BmKTX binding interface. Pharmacological experiments indicated that BmKTX-19 and BmKTX-196 peptides were specific inhibitors of the Kv1.3 channel and effectively suppressed cytokine secretion. In addition to the structural similarity between the designed and native peptides, both experimental alanine-scanning mutagenesis and computational simulation further indicated that the binding interface of wild-type BmKTX was successfully reoriented in BmKTX-19 and BmKTX-196, which adopted distinct toxin surfaces as binding interfaces. Together, these findings indicate not only the promising prospect of BmKTX-19 and BmKTX-196 as drug candidates but also the desirable feasibility of the evolution-guided peptide drug design for discovering numerous peptide drugs for the Kv1.3 channel.
Subject(s)
Amino Acids, Acidic/toxicity , Drug Design , Evolution, Molecular , Immunotherapy , Peptides/chemistry , Amino Acid Sequence , Cytokines/metabolism , HEK293 Cells , Humans , Kv1.3 Potassium Channel/antagonists & inhibitors , Kv1.3 Potassium Channel/chemistry , Models, Molecular , Molecular Sequence Data , Mutagenesis/drug effects , Potassium Channel Blockers/pharmacology , Protein Binding/drug effects , Scorpion Venoms/chemistry , Solutions , T-Lymphocytes/drug effects , T-Lymphocytes/metabolismABSTRACT
For elucidation of the structural and conformational requirements on the endotoxic and antagonistic activity of lipid A derivatives, we designed and synthesized lipid A analogues containing acidic amino acid residues in place of the non-reducing end phosphorylated glucosamine. Definite switching of the endotoxic or antagonistic activity was observed depending on the difference of the acidic groups (phosphoric acid or carboxylic acid) in the lipid A analogues.
