薬剤活性向上した大環状ペプチドを開発する新規戦略

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ABC-transporters are a large class of membrane bound proteins involved in the formation of drug resistance in eukaryotes and prokaryotes, especially with regard to anti cancer drugs and antibiotics. In collaboration with Prof. R. Tampe (University of Frankfurt), we have elicited de novo natural product-like peptides that interact with TmrAB - the closest homolog to the human transporter associated with antigen processing - using the RaPID selection platform. Selections were performed against the outward and inward facing conformation of TmrAB.Deep sequencing showed that after 6 rounds of selection the top 10 sequences accounted for >60% of the sequence reads. The top sequences were enriched up to 20-40% of the total population. The identified peptides where chemically synthesized and characterized by various assays. The peptides were shown to bind tightly to the protein with low nanomolar affinities (KD values are in the range of 2-30 nM).Characterization by S. Hank showed that some of the discovered peptides block TmrAB peptide translocation activity through inhibition of ATP hydrolysis. Interestingly, the peptides selected against the outward facing conformer were overall more efficient in blocking activity than the peptides selected against the inward conformer. As revealed by analytical size exclusion chromatography the V-D-Tyr 3 peptide binds most tightly to the TmrAB-ATP complex and was therefore chosen as a stabilizing ligand for crystallization and structure elucidation. Crystallization efforts are currently ongoing.