Effects of Tryptophan Content and Backbone Spacing on the Uptake Efficiency of Cell-Penetrating Peptides
Journal article, 2012

Cell-penetrating peptides (CPPs) are able to traverse cellular membranes and deliver macromolecular cargo. Uptake occurs through both endocytotic and nonendocytotic pathways, but the molecular requirements for efficient internalization are not fully understood. Here we investigate how the presence of tryptophans and their position within an oligoarginine influence uptake mechanism and efficiency. Flow cytometry and confocal fluorescence imaging are used to estimate uptake efficiency, intracellular distribution and toxicity in Chinese hamster ovarian cells. Further, membrane leakage and lipid membrane affinity are investigated. The peptides contain eight arginine residues and one to four tryptophans, the tryptophans positioned either at the N-terminus, in the middle, or evenly distributed along the amino acid sequence. Our data show that the intracellular distribution varies among peptides with different tryptophan content and backbone spacing. Uptake efficiency is higher for the peptides with four tryptophans in the middle, or evenly distributed along the peptide sequence, than for the peptide with four tryptophans at the N-terminus. All peptides display low cytotoxicity except for the one with four tryptophans at the N-terminus, which was moderately toxic. This finding is consistent with their inability to induce efficient leakage of dye from lipid vesicles. All peptides have comparable affinities for lipid vesicles, showing that lipid binding is not a decisive parameter for uptake. Our results indicate that tryptophan content and backbone spacing can affect both the CPP uptake efficiency and the CPP uptake mechanism. The low cytotoxicity of these peptides and the possibilities of tuning their uptake mechanism are interesting from a therapeutic point of view.

membrane-proteins

translocation

arginine-rich peptides

binding

mechanisms

delivery

endocytosis

plasma-membrane

internalization

molecular

transporters

Author

Hanna Rydberg

Chalmers, Chemical and Biological Engineering, Physical Chemistry

Maria Matson

Chalmers, Chemical and Biological Engineering, Physical Chemistry

Helene Åmand

Chalmers, Chemical and Biological Engineering, Physical Chemistry

Elin Esbjörner Winters

Chemical Biology

Bengt Nordén

Chalmers, Chemical and Biological Engineering, Physical Chemistry

Biochemistry

0006-2960 (ISSN) 1520-4995 (eISSN)

Vol. 51 27 5531-5539

Areas of Advance

Energy

Life Science Engineering (2010-2018)

Materials Science

Subject Categories

Chemical Sciences

DOI

10.1021/bi300454k

More information

Latest update

8/23/2021