Issue 4, 2017

A practical strategy to design and develop an isoform-specific fluorescent probe for a target enzyme: CYP1A1 as a case study

Abstract

The development of isoform-specific probe(s) for a target enzyme with multiple homologs is always challenging. Herein, a practical strategy was used to design and develop an isoform-specific probe for CYP1A1, a key cytochrome P450 isoenzyme involved in xenobiotic metabolism and bioactivation. On the basis of the subtle differences in 3D structure and substrate preference between CYP1A1 and its homolog CYP1A2, we proposed that it was possible to design a CYP1A1-specific probe via local modification of the reaction site on known CYP1A substrates. To validate this hypothesis, 4-hydroxy-1,8-naphthalimide (HN) was selected as the basic fluorophore due to its excellent optical properties, while a series of O-alkylated HN derivatives were prepared to evaluate their specificity towards CYP1A1. Our results revealed that the introduction of a chloroethyl to HN could get the best isoform selectivity towards CYP1A1 over other CYPs including CYP1A2. The newly developed probe NBCeN exhibited excellent specificity, high sensitivity, and a ratiometric fluorescence response following CYP1A1-catalyzed O-dechloroethylation. NBCeN was successfully used to real-time monitor the activity of CYP1A1 in complex biological samples and to rapidly screen CYP1A1 modulators in living systems. NBCeN could also be used for two-photon imaging of intracellular CYP1A1 in living cells and tissues with high ratiometric imaging resolution and deep tissue penetration. All these findings demonstrated that local modification of non-specific substrates was a practical strategy to develop an isoform-specific probe for a target isoenzyme, while NBCeN could serve as a specific imaging tool to explore the biological functions of CYP1A1 in complex biological systems.

Graphical abstract: A practical strategy to design and develop an isoform-specific fluorescent probe for a target enzyme: CYP1A1 as a case study

Supplementary files

Article information

Article type
Edge Article
Submitted
05 Sep 2016
Accepted
16 Dec 2016
First published
19 Dec 2016
This article is Open Access

All publication charges for this article have been paid for by the Royal Society of Chemistry
Creative Commons BY license

Chem. Sci., 2017,8, 2795-2803

A practical strategy to design and develop an isoform-specific fluorescent probe for a target enzyme: CYP1A1 as a case study

Z. Dai, L. Feng, Q. Jin, H. Cheng, Y. Li, J. Ning, Y. Yu, G. Ge, J. Cui and L. Yang, Chem. Sci., 2017, 8, 2795 DOI: 10.1039/C6SC03970G

This article is licensed under a Creative Commons Attribution 3.0 Unported Licence. You can use material from this article in other publications without requesting further permissions from the RSC, provided that the correct acknowledgement is given.

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