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Biosens Bioelectron. 2014 Nov 15;61:404-9. doi: 10.1016/j.bios.2014.05.031. Epub 2014 May 21.

Thymine-based molecular beacon for sensing adenosine based on the inhibition of S-adenosylhomocysteine hydrolase activity.

Author information

1
Department of Chemistry, National Sun Yat-sen University, 70, Lien-hai Road, Kaohsiung, Taiwan.
2
Department of Chemistry, National Sun Yat-sen University, 70, Lien-hai Road, Kaohsiung, Taiwan; School of Pharmacy, College of Pharmacy, Kaohsiung Medical University, Taiwan. Electronic address: tsengwl@mail.nsysu.edu.tw.

Abstract

This study presents a thymine (T)-based molecular beacon (MB) used for probing S-adenosylhomocysteine hydrolase (SAHH)-catalyzed hydrolysis of S-adenosylhomocysteine (SAH) and for sensing adenosine based on the inhibition of SAHH activity. The designed MB (T8-MB-T8) contained a 15-mer loop and a stem that consisted of a pair of 8-mer T bases, a fluorophore unit at the 5'-end, and a quencher unit at the 3'-end. In the presence of Hg(2+), a change in the conformation of T8-MB-T8 placed the fluorophore unit and the quencher in proximity to each other and caused collisional quenching of fluorescence between them. The Hg(2+)-induced fluorescence quenching of T8-MB-T8 occurred because the Hg(2+) induced T-T mismatches to form stable T-Hg(2+)-T coordination in the MB stem. SAHH catalyzed the hydrolysis of SAH to produce homocysteine. The generated homocysteine enabled the Hg(2+) to be removed from a hairpin-shaped T8-MB-T8 through the formation of a strong Hg(2+)-S bond, leading to the restoration of its fluorescence. The T8-MB-T8 · Hg(2+) probe showed a limit of detection for SAHH of 4 units L(-1) (approximately 0.24 nM) and was reusable for detecting the SAHH/SAH system. Because adenosine was an effective SAHH activity inhibitor, the T8-MB-T8 · Hg(2+) probe combining the SAHH and SAH systems was used for sensitive and selective detection of adenosine in urine without the interference of other adenosine analogs.

KEYWORDS:

Adenosine; Mercury ions; Molecular beacon; S-adenosylhomocysteine; S-adenosylhomocysteine hydrolase

PMID:
24914852
DOI:
10.1016/j.bios.2014.05.031
[Indexed for MEDLINE]

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