Membrane translocation of penetratin and its derivatives in different cell lines

T. Letoha, S. Gaá, C. Somlai, A. Czajlik, A. Perczel, B. Penke

Research output: Contribution to journalArticle

68 Citations (Scopus)

Abstract

The third helix of the homeodomain of the Antennapedia homeoprotein can translocate through the cell membrane into the nucleus and can be used as an intracellular vehicle for the delivery of oligopeptides and oligonucleotides. A 16-amino acid-long peptide fragment, called penetratin, is internalized by the cells in a specific, non-receptor-mediated manner. For a better understanding of the mechanism of the transfer, penetratin and two analogs were synthesized: (1) penetratin RQIKIWFQNRRMKWKK (peptide 1); (2) (6,14-Phe)-penetratin, RQIKIFFQNRRMKFKK (peptide 2); (3) dodecapeptide, RQIKIWF-R-KWKK (peptide 3); The conformation of penetratin peptides 1-3 was examined in both extracellular matrix-mimetic and membrane-mimetic environments. 1H-NMR and CD spectroscopic measurements were performed in mixtures of TFE/water with different ratios. Peptides 1-3 were labeled by reacting their N-terminal free amino group with fluorescein isothiocyanate (FITC). Membrane translocation of the labelled peptides was studied with cell cultures [WEHI 164 murine fibrosarcoma cells (WC/1); chicken fibroblast cells (CEC-32); chicken monocytic cells (HD-11); human fibroblast (SV 80) and human monocytic cells (MonoMac-6)]. Confocal laser scanning microscopy and flow cytometry assay were used to study membrane translocation. Amphiphilicity was calculated for each peptide. In our experiments all the penetratin peptides penetrated into the cells. Helical conformation and membrane translocation ability showed little correlation: substitution of the two Trp with Phe increased the stability of helical conformation but decreased membrane translocation activity. The results of fluorescence microscopy and flow cytometry show that penetratin can be translocated into the cells by two mechanisms: endocytosis and direct transport through the cell membrane.

Original languageEnglish
Pages (from-to)272-279
Number of pages8
JournalJournal of Molecular Recognition
Volume16
Issue number5
DOIs
Publication statusPublished - Sep 2003

Fingerprint

Peptides
Cells
Derivatives
Membranes
Cell Line
Conformations
Flow cytometry
Fibroblasts
Cell membranes
Chickens
Flow Cytometry
Laser Scanning Cytometry
Cell Membrane
Homeodomain Proteins
Oligopeptides
Peptide Fragments
Fibrosarcoma
Fluorescence microscopy
Polytetrafluoroethylene
Endocytosis

Keywords

  • Amphiphilicity
  • CD
  • Cell penetrating peptides
  • Conformation
  • FITC-labeling
  • NMR
  • Penetratin
  • Translocation mechanism

ASJC Scopus subject areas

  • Biochemistry
  • Genetics
  • Computer Vision and Pattern Recognition
  • Immunology
  • Molecular Biology

Cite this

Membrane translocation of penetratin and its derivatives in different cell lines. / Letoha, T.; Gaá, S.; Somlai, C.; Czajlik, A.; Perczel, A.; Penke, B.

In: Journal of Molecular Recognition, Vol. 16, No. 5, 09.2003, p. 272-279.

Research output: Contribution to journalArticle

@article{54af7c138f224aeaa8589c600a2ea2e2,
title = "Membrane translocation of penetratin and its derivatives in different cell lines",
abstract = "The third helix of the homeodomain of the Antennapedia homeoprotein can translocate through the cell membrane into the nucleus and can be used as an intracellular vehicle for the delivery of oligopeptides and oligonucleotides. A 16-amino acid-long peptide fragment, called penetratin, is internalized by the cells in a specific, non-receptor-mediated manner. For a better understanding of the mechanism of the transfer, penetratin and two analogs were synthesized: (1) penetratin RQIKIWFQNRRMKWKK (peptide 1); (2) (6,14-Phe)-penetratin, RQIKIFFQNRRMKFKK (peptide 2); (3) dodecapeptide, RQIKIWF-R-KWKK (peptide 3); The conformation of penetratin peptides 1-3 was examined in both extracellular matrix-mimetic and membrane-mimetic environments. 1H-NMR and CD spectroscopic measurements were performed in mixtures of TFE/water with different ratios. Peptides 1-3 were labeled by reacting their N-terminal free amino group with fluorescein isothiocyanate (FITC). Membrane translocation of the labelled peptides was studied with cell cultures [WEHI 164 murine fibrosarcoma cells (WC/1); chicken fibroblast cells (CEC-32); chicken monocytic cells (HD-11); human fibroblast (SV 80) and human monocytic cells (MonoMac-6)]. Confocal laser scanning microscopy and flow cytometry assay were used to study membrane translocation. Amphiphilicity was calculated for each peptide. In our experiments all the penetratin peptides penetrated into the cells. Helical conformation and membrane translocation ability showed little correlation: substitution of the two Trp with Phe increased the stability of helical conformation but decreased membrane translocation activity. The results of fluorescence microscopy and flow cytometry show that penetratin can be translocated into the cells by two mechanisms: endocytosis and direct transport through the cell membrane.",
keywords = "Amphiphilicity, CD, Cell penetrating peptides, Conformation, FITC-labeling, NMR, Penetratin, Translocation mechanism",
author = "T. Letoha and S. Ga{\'a} and C. Somlai and A. Czajlik and A. Perczel and B. Penke",
year = "2003",
month = "9",
doi = "10.1002/jmr.637",
language = "English",
volume = "16",
pages = "272--279",
journal = "Journal of Molecular Recognition",
issn = "0952-3499",
publisher = "John Wiley and Sons Ltd",
number = "5",

}

TY - JOUR

T1 - Membrane translocation of penetratin and its derivatives in different cell lines

AU - Letoha, T.

AU - Gaá, S.

AU - Somlai, C.

AU - Czajlik, A.

AU - Perczel, A.

AU - Penke, B.

PY - 2003/9

Y1 - 2003/9

N2 - The third helix of the homeodomain of the Antennapedia homeoprotein can translocate through the cell membrane into the nucleus and can be used as an intracellular vehicle for the delivery of oligopeptides and oligonucleotides. A 16-amino acid-long peptide fragment, called penetratin, is internalized by the cells in a specific, non-receptor-mediated manner. For a better understanding of the mechanism of the transfer, penetratin and two analogs were synthesized: (1) penetratin RQIKIWFQNRRMKWKK (peptide 1); (2) (6,14-Phe)-penetratin, RQIKIFFQNRRMKFKK (peptide 2); (3) dodecapeptide, RQIKIWF-R-KWKK (peptide 3); The conformation of penetratin peptides 1-3 was examined in both extracellular matrix-mimetic and membrane-mimetic environments. 1H-NMR and CD spectroscopic measurements were performed in mixtures of TFE/water with different ratios. Peptides 1-3 were labeled by reacting their N-terminal free amino group with fluorescein isothiocyanate (FITC). Membrane translocation of the labelled peptides was studied with cell cultures [WEHI 164 murine fibrosarcoma cells (WC/1); chicken fibroblast cells (CEC-32); chicken monocytic cells (HD-11); human fibroblast (SV 80) and human monocytic cells (MonoMac-6)]. Confocal laser scanning microscopy and flow cytometry assay were used to study membrane translocation. Amphiphilicity was calculated for each peptide. In our experiments all the penetratin peptides penetrated into the cells. Helical conformation and membrane translocation ability showed little correlation: substitution of the two Trp with Phe increased the stability of helical conformation but decreased membrane translocation activity. The results of fluorescence microscopy and flow cytometry show that penetratin can be translocated into the cells by two mechanisms: endocytosis and direct transport through the cell membrane.

AB - The third helix of the homeodomain of the Antennapedia homeoprotein can translocate through the cell membrane into the nucleus and can be used as an intracellular vehicle for the delivery of oligopeptides and oligonucleotides. A 16-amino acid-long peptide fragment, called penetratin, is internalized by the cells in a specific, non-receptor-mediated manner. For a better understanding of the mechanism of the transfer, penetratin and two analogs were synthesized: (1) penetratin RQIKIWFQNRRMKWKK (peptide 1); (2) (6,14-Phe)-penetratin, RQIKIFFQNRRMKFKK (peptide 2); (3) dodecapeptide, RQIKIWF-R-KWKK (peptide 3); The conformation of penetratin peptides 1-3 was examined in both extracellular matrix-mimetic and membrane-mimetic environments. 1H-NMR and CD spectroscopic measurements were performed in mixtures of TFE/water with different ratios. Peptides 1-3 were labeled by reacting their N-terminal free amino group with fluorescein isothiocyanate (FITC). Membrane translocation of the labelled peptides was studied with cell cultures [WEHI 164 murine fibrosarcoma cells (WC/1); chicken fibroblast cells (CEC-32); chicken monocytic cells (HD-11); human fibroblast (SV 80) and human monocytic cells (MonoMac-6)]. Confocal laser scanning microscopy and flow cytometry assay were used to study membrane translocation. Amphiphilicity was calculated for each peptide. In our experiments all the penetratin peptides penetrated into the cells. Helical conformation and membrane translocation ability showed little correlation: substitution of the two Trp with Phe increased the stability of helical conformation but decreased membrane translocation activity. The results of fluorescence microscopy and flow cytometry show that penetratin can be translocated into the cells by two mechanisms: endocytosis and direct transport through the cell membrane.

KW - Amphiphilicity

KW - CD

KW - Cell penetrating peptides

KW - Conformation

KW - FITC-labeling

KW - NMR

KW - Penetratin

KW - Translocation mechanism

UR - http://www.scopus.com/inward/record.url?scp=0141942111&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0141942111&partnerID=8YFLogxK

U2 - 10.1002/jmr.637

DO - 10.1002/jmr.637

M3 - Article

C2 - 14523940

AN - SCOPUS:0141942111

VL - 16

SP - 272

EP - 279

JO - Journal of Molecular Recognition

JF - Journal of Molecular Recognition

SN - 0952-3499

IS - 5

ER -