title: An unprecedented alteration in mode of action of IsCT resulting its translocation into bacterial cytoplasm and inhibition of macromolecular syntheses.
abstract: IsCT, a 13-residue, non-cell-selective antimicrobial peptide is comprised of mostly hydrophobic residues and lesser cationic residues. Assuming that placement of an additional positive charge in the non-polar face of IsCT could reduce its hydrophobic interaction, resulting in its reduction of cytotoxicity, an analog, I9K-IsCT was designed. Two more analogs, namely, E7K-IsCT and E7K,I9K-IsCT, were designed to investigate the impact of positive charges in the polar face as well as polar and non-polar faces at a time. These amino acid substitutions resulted in a significant enhancement of therapeutic potential of IsCT. IsCT and E7K-IsCT seem to target bacterial membrane for their anti-bacterial activity. However, I9K-IsCT and E7K,I9K-IsCT inhibited nucleic acid and protein syntheses in tested E. coli without perturbing its membrane. This was further supported by the observation that NBD-IsCT localized onto bacterial membrane while NBD-labeled I9K-IsCT and E7K,I9K-IsCT translocated into bacterial cytoplasm. Interestingly, IsCT and E7K-IsCT were significantly helical while I9K-IsCT and E7K,I9K-IsCT were mostly unstructured with no helix content in presence of mammalian and bacterial membrane-mimetic lipid vesicles. Altogether, the results identify two novel cell-selective analogs of IsCT with new prototype amino acid sequences that can translocate into bacterial cytoplasm without any helical structure and inhibit macromolecular syntheses.
authors.affiliation: Molecular and Structural Biology Division, CSIR-Central Drug Research Institute, Sector 10, Jankipuram Extension, Sitapur Road, Lucknow-226 031, India.
authors.forename: Jitendra K
authors.initials: JK
authors.lastname: Tripathi
authors.affiliation: Electron Microscopy Unit, CSIR-Central Drug Research Institute, Sector 10, Jankipuram Extension, Sitapur Road, Lucknow-226 031, India.
authors.forename: Manoj
authors.initials: M
authors.lastname: Kathuria
authors.affiliation: Molecular and Structural Biology Division, CSIR-Central Drug Research Institute, Sector 10, Jankipuram Extension, Sitapur Road, Lucknow-226 031, India.
authors.forename: Amit
authors.initials: A
authors.lastname: Kumar
authors.affiliation: Electron Microscopy Unit, CSIR-Central Drug Research Institute, Sector 10, Jankipuram Extension, Sitapur Road, Lucknow-226 031, India.
authors.forename: Kalyan
authors.initials: K
authors.lastname: Mitra
authors.affiliation: Molecular and Structural Biology Division, CSIR-Central Drug Research Institute, Sector 10, Jankipuram Extension, Sitapur Road, Lucknow-226 031, India.
authors.forename: Jimut K
authors.initials: JK
authors.lastname: Ghosh
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D023181:Antimicrobial Cationic Peptides / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology*; D001419:Bacteria / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D003593:Cytoplasm / Q000378:metabolism; D004912:Erythrocytes / Q000187:drug effects; D004926:Escherichia coli / Q000187:drug effects / Q000378:metabolism; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D051379:Mice; D008826:Microbial Sensitivity Tests; D050296:Microbial Viability / Q000187:drug effects; D010539:Permeability; D021381:Protein Transport
chemical_list: D000900:Anti-Bacterial Agents; D023181:Antimicrobial Cationic Peptides