Change search
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf
Modeling the endosomal escape of cell-penetrating peptides using a transmembrane pH gradient
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Stockholm University, Faculty of Science, Department of Neurochemistry.
Stockholm University, Faculty of Science, Department of Neurochemistry.ORCID iD: 0000-0001-7746-8574
Show others and affiliations
2013 (English)In: Biochimica et Biophysica Acta - Biomembranes, ISSN 0005-2736, E-ISSN 1879-2642, Vol. 1828, no 4, 1198-1204 p.Article in journal (Refereed) Published
Abstract [en]

Cell-penetrating peptides (CPPs) can internalize into cells with covalently or non-covalently bound biologically active cargo molecules, which by themselves are not able to pass the cell membrane. Direct penetration and endocytosis are two main pathways suggested for the cellular uptake of CPPs. Cargo molecules which have entered the cell via an endocytotic pathway must be released from the endosome before degradation by enzymatic processes and endosomal acidification. Endosomal entrapment seems to be a major limitation in delivery of these molecules into the cytoplasm. Bacteriorhodopsin (BR) asymmetrically introduced into large unilamellar vesicles (LUVs) was used to induce a pH gradient across the lipid bilayer. By measuring pH outside the LUVs, we observed light-induced proton pumping mediated by BR from the outside to the inside of the LUVs, creating an acidic pH inside the LUVs, similar to the late endosomes in vivo. Here we studied the background mechanism(s) of endosomal escape. 20% negatively charged LUVs were used as model endosomes with incorporated BR into the membrane and fluorescein-labeled CPPs entrapped inside the LUVs, together with a fluorescence quencher. The translocation of different CPPs in the presence of a pH gradient across the membrane was studied. The results show that the light-induced pH gradient induced by BR facilitates vesicle membrane translocation, particularly for the intermediately hydrophobic CPPs, and much less for hydrophilic CPPs. The presence of chloroquine inside the LUVs or addition of pyrenebutyrate outside the LUVs destabilizes the vesicle membrane, resulting in significant changes of the pH gradient across the membrane.

Place, publisher, year, edition, pages
2013. Vol. 1828, no 4, 1198-1204 p.
Keyword [en]
Cell-penetrating peptide, Endosomal escape, Bacteriorhodopsin, Large unilamellar vesicle, Fluorescein-label, Membrane translocation
National Category
Biophysics Chemical Sciences
Research subject
Biophysics
Identifiers
URN: urn:nbn:se:su:diva-84928DOI: 10.1016/j.bbamem.2012.12.008ISI: 000316522100003PubMedID: 23261392OAI: oai:DiVA.org:su-84928DiVA: diva2:581960
Funder
Swedish Research CouncilVINNOVASwedish Foundation for Strategic Research
Available from: 2013-01-03 Created: 2013-01-03 Last updated: 2017-12-06Bibliographically approved
In thesis
1. Biophysical studies of peptides with functions in biotechnology and biology
Open this publication in new window or tab >>Biophysical studies of peptides with functions in biotechnology and biology
2012 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

My thesis concerns spectroscopic studies (NMR, CD and fluorescence) of peptides with functions in biotechnology and biology, and their interactions with a model membrane (large unilamellar phospholipid vesicles).

The resorufin-based arsenical hairpin binder (ReAsH) bound to a short peptide is a useful fluorescent tag for genetic labeling of proteins in living cells. A hairpin structure with some resemblance to type II β-turn was determined by NMR structure calculations (Paper I).

Cell-penetrating peptides (CPPs) are short (30-35 residues), often rich in basic amino acids such as Arg. They can pass through the cell membrane and deliver bioactive cargoes, making them useful for biotechnical and pharmacological applications. The mechanisms of cellular uptake and membrane translocation are under debate. Understanding the mechanistic aspects of CPPs is the major focus of Papers II, III, and IV.

The effect of the pyrenebutyrate (PB) on the cellular uptake, membrane translocation and perturbation of several CPPs from different subgroups was investigated (Paper II). We concluded that both charge and hydrophobicity of the CPP affect the cellular uptake and membrane translocation efficiency.

Endosomal escape is a crucial challenge for the CPP applications. We modeled the endosome and endosomal escape for different CPPs to investigate the corresponding molecular mechanisms (Papers III and IV). Hydrophobic CPPs were able to translocate across the model membrane in the presence of a pH gradient, produced by bacteriorhodopsin proton pumping, whereas a smaller effect was observed for hydrophilic CPPs.

Dynorphin A (Dyn A) peptide mutations are associated with neurodegenerative disorders, without involvement of the opioid receptors. The non-opioid activities of Dyn A may involve membrane perturbations. Model membrane-perturbations by three Dyn A mutants were investigated (Paper V). The results showed effects to different degrees largely in accordance with their neurotoxic effects.

Place, publisher, year, edition, pages
Stockholm: Department of Biochemistry and Biophysics, Stockholm University, 2012. 75 p.
Keyword
Genetic fluorescence label, Biarsenical tetracysteine motif, Cell-penetrating peptides, Large unilamellar vesicles, Pyrenebutyrate, Endosomal escape, Membrane perturbation, Bacteriorhodopsin, Dynorphin
National Category
Biophysics
Research subject
Biophysics
Identifiers
urn:nbn:se:su:diva-66948 (URN)978-91-7447-417-6 (ISBN)
Public defence
2012-02-14, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 10:00 (English)
Opponent
Supervisors
Note

At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 4: Manuscript.

Available from: 2012-01-23 Created: 2011-12-22 Last updated: 2013-04-09Bibliographically approved

Open Access in DiVA

No full text

Other links

Publisher's full textPubMed

Search in DiVA

By author/editor
Madani, FatemehAbdo, RaniaLindberg, StaffanLangel, ÜloGräslund, Astrid
By organisation
Department of Biochemistry and BiophysicsDepartment of Neurochemistry
In the same journal
Biochimica et Biophysica Acta - Biomembranes
BiophysicsChemical Sciences

Search outside of DiVA

GoogleGoogle Scholar

doi
pubmed
urn-nbn

Altmetric score

doi
pubmed
urn-nbn
Total: 105 hits
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf