RRC ID 69059
Author Suzuki M, Iwaki K, Kikuchi M, Fujiwara K, Doi N.
Title Characterization of the membrane penetration-enhancing peptide S19 derived from human syncytin-1 for the intracellular delivery of TAT-fused proteins.
Journal Biochem Biophys Res Commun
Abstract Although cell-penetrating peptides such as the HIV-derived TAT peptide have been used as tools for the intracellular delivery of therapeutic peptides and proteins, a problem persists: the endosomal escape efficiency is low. Previously, we found that the fusogenic peptide S19, derived from the human protein syncytin-1, enhance the endosomal escape efficiency of proteins that incorporated by endocytosis via TAT. In this study, we first performed Ala-scanning mutagenesis of S19, and found that all Ile, Val, Leu and Phe with high β-sheet forming propensities in S19 are important for the intracellular uptake of S19-TAT-fused proteins. In a secondary structure analysis of the mutated S19-TAT peptides in the presence of liposomes mimicking late endosomes (LEs), the CD spectra of V3A and I4A mutants with low uptake activity showed the appearance of an α-helix structure, whereas the mutant G5A retained both the uptake activity and the β-structure. In addition, we investigated the appropriate linking position and order of the S19 and TAT peptides to a cargo protein including an apoptosis-induced peptide and found that both the previous C-terminal S19-TAT tag and the N-terminal TAT-S19 tag promote the cytoplasmic delivery of the fusion protein. These results and previous results suggest that the interaction of TAT with the LE membrane causes a structural change in S19 from a random coil to a β-strand and that the subsequent parallel β-sheet formation between two S19 peptides may promote adjacent TAT dimerization, resulting in endosomal escape from the LE membrane.
Volume 586
Pages 63-67
Published 2022-1-1
DOI 10.1016/j.bbrc.2021.11.065
PII S0006-291X(21)01582-5
PMID 34826702
MeSH Amino Acid Substitution Cell Line, Tumor Cell Membrane / metabolism* Cell Membrane Permeability Endosomes / chemistry Endosomes / metabolism Gene Expression Gene Products, env / genetics Gene Products, env / metabolism* Gene Products, tat / genetics Gene Products, tat / metabolism* Genes, Reporter Green Fluorescent Proteins / genetics Green Fluorescent Proteins / metabolism HeLa Cells Humans Liposomes / chemistry Liposomes / metabolism Optical Imaging Peptides / genetics Peptides / metabolism* Plasmids / chemistry Plasmids / metabolism* Pregnancy Proteins / genetics Pregnancy Proteins / metabolism* Protein Conformation, alpha-Helical Protein Conformation, beta-Strand Protein Transport Recombinant Fusion Proteins / genetics Recombinant Fusion Proteins / metabolism* Transduction, Genetic
IF 2.985
Human and Animal Cells DU145(RCB2143) HeLa(RCB0007)