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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.

SUBMITTER: Tripathi JK

PROVIDER: S-EPMC4360471 | biostudies-literature | 2015 Mar

REPOSITORIES: biostudies-literature

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An unprecedented alteration in mode of action of IsCT resulting its translocation into bacterial cytoplasm and inhibition of macromolecular syntheses.

Tripathi Jitendra K JK Kathuria Manoj M Kumar Amit A Kumar Amit A Mitra Kalyan K Ghosh Jimut K JK

Scientific reports 20150316

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-po ...[more]

PMID: 25773522

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Project description:IntroductionFrom viruses to organelles, fusion of biological membranes is used by diverse biological systems to deliver macromolecules across membrane barriers. Membrane fusion is also a potentially efficient mechanism for the delivery of macromolecular therapeutics to the cellular cytoplasm. However, a key shortcoming of existing fusogenic liposomal systems is that they are inefficient, requiring a high concentration of fusion-promoting lipids in order to cross cellular membrane barriers.ObjectivesToward addressing this limitation, our experiments explore the extent to which membrane fusion can be amplified by using the process of lipid membrane phase separation to concentrate fusion-promoting lipids within distinct regions of the membrane surface.MethodsWe used confocal fluorescence microscopy to investigate the integration of fusion-promoting lipids into a ternary lipid membrane system that separated into liquid-ordered and liquid-disordered membrane phases. Additionally, we quantified the impact of membrane phase separation on the efficiency with which liposomes transferred lipids and encapsulated macromolecules to cells, using a combination of confocal fluorescence imaging and flow cytometry.ResultsHere we report that concentrating fusion-promoting lipids within phase-separated lipid domains on the surfaces of liposomes significantly increases the efficiency of liposome fusion with model membranes and cells. In particular, membrane phase separation enhanced the delivery of lipids and model macromolecules to the cytoplasm of tumor cells by at least 4-fold in comparison to homogenous liposomes.ConclusionsOur findings demonstrate that phase separation can enhance membrane fusion by locally concentrating fusion-promoting lipids on the surface of liposomes. This work represents the first application of lipid membrane phase separation in the design of biomaterials-based delivery systems. Additionally, these results lay the ground work for developing fusogenic liposomes that are triggered by physical and molecular cues associated with target cells.

Dataset Information

An unprecedented alteration in mode of action of IsCT resulting its translocation into bacterial cytoplasm and inhibition of macromolecular syntheses.

Publications

An unprecedented alteration in mode of action of IsCT resulting its translocation into bacterial cytoplasm and inhibition of macromolecular syntheses.

Similar Datasets

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