G-Quadruplex-Forming Oligonucleotide Conjugated to Magnetic Nanoparticles: Synthesis, Characterization, and Enzymatic Stability Assays

In the present work, we report the conjugation of superparamagnetic nanoparticles to a fluorescently labeled oligodeoxyribonucleotide (ODN) able to fold into stable unimolecular guanine quadruple helix under proper ion conditions by means of its thrombin-binding aptamer (TBA) sequence. The novel mod...

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Bibliographic Details
Published in:Bioconjugate chemistry 2012-03, Vol.23 (3), p.382-391
Main Authors: Musumeci, Domenica, Oliviero, Giorgia, Roviello, Giovanni N, Bucci, Enrico M, Piccialli, Gennaro
Format: Article
Language:English
Subjects:
Chromatography, High Pressure Liquid
Enzyme Stability
Enzymes
Fluorescence
G-Quadruplexes
Ions
Magnetics
Nanoparticles
Oligonucleotides - chemistry
Spectrum analysis
Spectrum Analysis - methods
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Summary:In the present work, we report the conjugation of superparamagnetic nanoparticles to a fluorescently labeled oligodeoxyribonucleotide (ODN) able to fold into stable unimolecular guanine quadruple helix under proper ion conditions by means of its thrombin-binding aptamer (TBA) sequence. The novel modified ODN, which contained a fluorescent dUPy unit at 3′-end and a 12-amino-dodecyl spacer (C12–NH2) at 5′ terminus, was characterized by ESI-MS and optical spectroscopy (UV, CD, fluorescence), and analyzed by RP-HPLC chromatography and electrophoresis. From CD and fluorescence experiments, we verified that dUPy and C12–NH2 incorporation does not interfere with the conformational stability of the G-quadruplex. Subsequently, the conjugation of the pyrene-labeled ODN with the magnetite particles was performed, and the ODN-conjugated nanoparticles were studied through optical spectroscopy (UV, CD, fluorescence) and by enzymatic and chemical assays. We found that the nanoparticles enhanced the stability of the TBA ODN to enzymatic degradation. Finally, we evaluated the amount of the TBA-conjugated nanoparticles immobilized on a magnetic separator in view of the potential use of the nanosystem for the magnetic capture of thrombin from complex mixtures.
ISSN:1043-1802
1520-4812
DOI:10.1021/bc200305t