Limits...
Efficient Synthesis of Peptide and Protein Functionalized Pyrrole-Imidazole Polyamides Using Native Chemical Ligation.

Janssen BM, van Ommeren SP, Merkx M - Int J Mol Sci (2015)

Bottom Line: The effect of Py-Im polyamide conjugation on DNA binding was investigated by Surface Plasmon Resonance (SPR).Although the synthesis of different protein-Py-Im-polyamide conjugates was successful, attenuation of DNA affinity was observed, in particular for the protein-Py-Im-polyamide conjugates.The practical use of protein-Py-Im-polyamide conjugates for addressing DNA structures in an orthogonal but non-covalent manner, therefore, remains to be established.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Chemical Biology and Institute for Complex Molecular Systems, Eindhoven University of Technology, Den Dolech 2, 5600 MB Eindhoven, The Netherlands. b.m.g.janssen@tue.nl.

ABSTRACT
The advancement of DNA-based bionanotechnology requires efficient strategies to functionalize DNA nanostructures in a specific manner with other biomolecules, most importantly peptides and proteins. Common DNA-functionalization methods rely on laborious and covalent conjugation between DNA and proteins or peptides. Pyrrole-imidazole (Py-Im) polyamides, based on natural minor groove DNA-binding small molecules, can bind to DNA in a sequence specific fashion. In this study, we explore the use of Py-Im polyamides for addressing proteins and peptides to DNA in a sequence specific and non-covalent manner. A generic synthetic approach based on native chemical ligation was established that allows efficient conjugation of both peptides and recombinant proteins to Py-Im polyamides. The effect of Py-Im polyamide conjugation on DNA binding was investigated by Surface Plasmon Resonance (SPR). Although the synthesis of different protein-Py-Im-polyamide conjugates was successful, attenuation of DNA affinity was observed, in particular for the protein-Py-Im-polyamide conjugates. The practical use of protein-Py-Im-polyamide conjugates for addressing DNA structures in an orthogonal but non-covalent manner, therefore, remains to be established.

No MeSH data available.


Related in: MedlinePlus

Synthetic scheme for the preparation of Py–Im polyamide bearing a PEGylated and tert-butylthiol protected cysteine for native chemical ligation purposes. The tert-butylthiol protected cysteine can be introduced at the C-terminus (a) or N-terminus (b) of the Py–Im polyamide. Introduction at the C-terminus requires cleavage from the resin using N,N-bis(aminopropyl)methylamine (BDp) followed by conjugation of the pre-synthesized PEG3-Cys(StBu) to the free amine of BDp. Introduction of PEGylated and tert-butylthiol protected cystein at the N-terminus can be performed on resin followed by cleavage from the resin using 3-(dimethylamino)-propylamine (Dp).
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4490465&req=5

ijms-16-12631-f006: Synthetic scheme for the preparation of Py–Im polyamide bearing a PEGylated and tert-butylthiol protected cysteine for native chemical ligation purposes. The tert-butylthiol protected cysteine can be introduced at the C-terminus (a) or N-terminus (b) of the Py–Im polyamide. Introduction at the C-terminus requires cleavage from the resin using N,N-bis(aminopropyl)methylamine (BDp) followed by conjugation of the pre-synthesized PEG3-Cys(StBu) to the free amine of BDp. Introduction of PEGylated and tert-butylthiol protected cystein at the N-terminus can be performed on resin followed by cleavage from the resin using 3-(dimethylamino)-propylamine (Dp).


Efficient Synthesis of Peptide and Protein Functionalized Pyrrole-Imidazole Polyamides Using Native Chemical Ligation.

Janssen BM, van Ommeren SP, Merkx M - Int J Mol Sci (2015)

Synthetic scheme for the preparation of Py–Im polyamide bearing a PEGylated and tert-butylthiol protected cysteine for native chemical ligation purposes. The tert-butylthiol protected cysteine can be introduced at the C-terminus (a) or N-terminus (b) of the Py–Im polyamide. Introduction at the C-terminus requires cleavage from the resin using N,N-bis(aminopropyl)methylamine (BDp) followed by conjugation of the pre-synthesized PEG3-Cys(StBu) to the free amine of BDp. Introduction of PEGylated and tert-butylthiol protected cystein at the N-terminus can be performed on resin followed by cleavage from the resin using 3-(dimethylamino)-propylamine (Dp).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4490465&req=5

ijms-16-12631-f006: Synthetic scheme for the preparation of Py–Im polyamide bearing a PEGylated and tert-butylthiol protected cysteine for native chemical ligation purposes. The tert-butylthiol protected cysteine can be introduced at the C-terminus (a) or N-terminus (b) of the Py–Im polyamide. Introduction at the C-terminus requires cleavage from the resin using N,N-bis(aminopropyl)methylamine (BDp) followed by conjugation of the pre-synthesized PEG3-Cys(StBu) to the free amine of BDp. Introduction of PEGylated and tert-butylthiol protected cystein at the N-terminus can be performed on resin followed by cleavage from the resin using 3-(dimethylamino)-propylamine (Dp).
Bottom Line: The effect of Py-Im polyamide conjugation on DNA binding was investigated by Surface Plasmon Resonance (SPR).Although the synthesis of different protein-Py-Im-polyamide conjugates was successful, attenuation of DNA affinity was observed, in particular for the protein-Py-Im-polyamide conjugates.The practical use of protein-Py-Im-polyamide conjugates for addressing DNA structures in an orthogonal but non-covalent manner, therefore, remains to be established.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Chemical Biology and Institute for Complex Molecular Systems, Eindhoven University of Technology, Den Dolech 2, 5600 MB Eindhoven, The Netherlands. b.m.g.janssen@tue.nl.

ABSTRACT
The advancement of DNA-based bionanotechnology requires efficient strategies to functionalize DNA nanostructures in a specific manner with other biomolecules, most importantly peptides and proteins. Common DNA-functionalization methods rely on laborious and covalent conjugation between DNA and proteins or peptides. Pyrrole-imidazole (Py-Im) polyamides, based on natural minor groove DNA-binding small molecules, can bind to DNA in a sequence specific fashion. In this study, we explore the use of Py-Im polyamides for addressing proteins and peptides to DNA in a sequence specific and non-covalent manner. A generic synthetic approach based on native chemical ligation was established that allows efficient conjugation of both peptides and recombinant proteins to Py-Im polyamides. The effect of Py-Im polyamide conjugation on DNA binding was investigated by Surface Plasmon Resonance (SPR). Although the synthesis of different protein-Py-Im-polyamide conjugates was successful, attenuation of DNA affinity was observed, in particular for the protein-Py-Im-polyamide conjugates. The practical use of protein-Py-Im-polyamide conjugates for addressing DNA structures in an orthogonal but non-covalent manner, therefore, remains to be established.

No MeSH data available.


Related in: MedlinePlus