Fluorophore-Dependent Cleavage Of Disulfide Bond Leading To A Highly Selective Fluorescent Probe Of Thioredoxin

Thioredoxin(Trx)system is one of the important systems regulating the intracellular redox signal and maintaining the redox homeostasis.This system with indispensable significance in physiology and pathology is closely related to many human diseases such as neurodegenerative diseases,diabetes and cancers.Moreover,Trx is also a potential target for the development of drugs inhibiting the growth of cancer cells.Therefore,exploring some methods of specifically detecting Trx is critical to understand its function in physiological and pathological processes.Small molecule fluorescent probes with appropriate specificity and higher spatiotemporal resolution have received more attention.So far,the fluorophores with short emission wavelength of the probes used to detect Trx,which limits their further biological applications.It is necessary to explore some fluorescent probes with longer wavelength,such as near-infrared fluorophores,that specifically detect Trx.In this paper,we designed and screened a near-infrared fluorescent probe quickly responding to Trx based on the reducibility of Trx to disulfide bonds.Next,we explored the optical properties,stability and biological application of this probe in subsequent tests.The contents are as following:1.Firstly,the structure characteristics of known mammalian Trx family members and their physiological functions are briefly introduced.And we summarized the development of traditional methods of detecting the activity of Trx,and then introduced the progresses of fluorescence methods of detecting the redox state of Trx.In addition,the development of disulfide-based fluorescent probes is briefly reviewed,and then the near-infrared fluorescent probes for recogniting biothiol and selenol are introduced.2.The in vitro models of Trx and GSH system were used to screen the reactivity of five symmetrical chain-disulfide compounds with Trx and GSH,respectively.And the bis-hydroxyethyl disulfide(HEDS)with a better reactivity to Trx was obtained.Next,using the HEDS as the recognition site of probe and selecting different fluorophores,we designed and synthesized four potential probes for Trx.And the near-infrared probe(NBL-SS)was screened by the reaction kinetic models.Moreover,the related in vitro tests of NBL-SS with Trx were performed and the probe was successfully applied to the detection for the activity of Trx in cells and zebrafish,and then we explored the feasibility of the fluorescent method of detecting the activity of Trx in cell lysates by using the fluorescence change of NBL-SS.