ZHAO H (2019).Pyrofosfataasien havaitseminen fluoresoivalla menetelmllä.

https://www.ncbi.nlm.nih.gov/pubmed/30572018

Mol Cell Probes. 2019 Feb;43:29-33. doi: 10.1016/j.mcp.2018.12.003. Epub 2018 Dec 17.

A novel fluorometric method for inorganic pyrophosphatase detection based on G-quadruplex-thioflavin T.

Zhao H¹, Ma C², Chen M¹.

1

School of Life Sciences, Central South University, Changsha, 410013, China.

2

School of Life Sciences, Central South University, Changsha, 410013, China. Electronic address: macb2012@csu.edu.cn.

Abstract

In this paper, we propose a fluorometric approach for the highly sensitive detection of inorganic pyrophosphatase (PPase) based on G-quadruplex-thioflavin T (ThT). In the absence of PPase, Cu²⁺ can coordinate with pyrophosphate (PPi) to generate a Cu²⁺/PPi complex. Then the G-rich sequence folds into the G-quadruplex structure, which can combine with ThT to generate a remarkable fluorescent signal. In the presence of PPase, the coordinated compound can be destroyed by the PPase catalyzed hydrolysis of PPi into inorganic phosphate (Pi). The subsequent release of Cu²⁺ can compete with ThT to induce a tighter G-quadruplex structure, causing the release of ThT and a sharp fluorescence decrease. Based on this mechanism, a facile and quantitative strategy for PPase detection was developed. The fluorescence intensity of the system shows a linear relationship with the PPase activities in the range of 0.5-30 U/L with a detection limit as low as 0.48 U/L. The proposed strategy for fluorescence spectrometric PPase detection is convenient, cost effective, and sensitive. This can be utilized to evaluate the inhibition effect of NaF on PPase as well as diagnose PPase-related diseases.

Copyright © 2018 Elsevier Ltd. All rights reserved.

KEYWORDS:

Fluorescence; G-quadruplex-thioflavin T; Label-free; Pyrophosphatase

PMID:

30572018

DOI:

10.1016/j.mcp.2018.12.003

[Indexed for MEDLINE]