Estrogen sulfotransferase(SULT1E1),an essential conjugative enzyme in mammals,plays a crucial role in both estrogen homeostasis and xenobiotic metabolism.Deciphering the dynamic changes in SULT1E1function under specif...Estrogen sulfotransferase(SULT1E1),an essential conjugative enzyme in mammals,plays a crucial role in both estrogen homeostasis and xenobiotic metabolism.Deciphering the dynamic changes in SULT1E1function under specific physiological or pathological conditions and discovering SULT1E1 modulators require practical and highly efficient tools for sensing SULT1E1 in biological context.Herein,we showcase a scaffold-seeking and structural optimization strategy for the rational engineering of isoform-specific fluorescent substrates for SULT1E1.First,docking-based virtual screening coupled with biochemical assays suggested that N–butyl–4-hydroxyphenyl-1,8-naphthalimide(HPN) was a suitable scaffold for constructing the fluorescent substrates for SULT1E1,but this fluorophore could be metabolized by multiple SULT isoforms.To develop isoform-specific substrates for SULT1E1,various substituents were introduced on the north part of HPN to explore the structure-enzyme specificity relationships of HPN derivatives as SULT1E1substrates.After molecular docking and experimental validation,an isoform-specific fluorescent substrate(HPN10) for SULT1E1 was successfully engineered.HPN10 demonstrated exceptional isoform-specificity,ultra-high sensitivity,and favorable signal-to-noise ratio(212).HPN10 excelled in the precise sensing of SULT1E1 activities in complex biological matrices,including cellular specimens and liver preparations.HPN10 immensely facilitated the discovery and characterization of SULT1E1 inhibitors,while tetrabromobisphenol A(TBBPA,half inhibitory concentration(IC_(50)) = 31.5 ± 3.4 nmol/L) was identified as a potent SULT1E1 inhibitor that could strongly block SULT1E1 activities in living cells.Collectively,this work presents a practical and efficient strategy for the rational engineering of isoform-specific fluorescent substrates for target conjugative enzyme(s),while HPN10 emerges as a reliable SULT1E1-activatable tool for functional sensing and drug discovery.展开更多
基金supported by Natural Science Foundation of China (Nos.U23A20516,82273897,81922070)Organizational Key Research and Development Program of Shanghai University of Traditional Chinese Medicine (No.2023YZZ02)+3 种基金the State Key Laboratory of Fine Chemicals,Dalian University of Technology (Nos.KF2202,KF2414)Shanghai Municipal Health Commission’s TCM research project (No.2022CX005)Future Plan for Traditional Chinese Medicine development of Science and Technology of Shanghai Municipal Hospital of TCM (Nos.WL-XJRY-2021010 K,WLGNDBZPY-2022001 K)Shanghai Jing’an District Health Commission (No.2024QT03)。
文摘Estrogen sulfotransferase(SULT1E1),an essential conjugative enzyme in mammals,plays a crucial role in both estrogen homeostasis and xenobiotic metabolism.Deciphering the dynamic changes in SULT1E1function under specific physiological or pathological conditions and discovering SULT1E1 modulators require practical and highly efficient tools for sensing SULT1E1 in biological context.Herein,we showcase a scaffold-seeking and structural optimization strategy for the rational engineering of isoform-specific fluorescent substrates for SULT1E1.First,docking-based virtual screening coupled with biochemical assays suggested that N–butyl–4-hydroxyphenyl-1,8-naphthalimide(HPN) was a suitable scaffold for constructing the fluorescent substrates for SULT1E1,but this fluorophore could be metabolized by multiple SULT isoforms.To develop isoform-specific substrates for SULT1E1,various substituents were introduced on the north part of HPN to explore the structure-enzyme specificity relationships of HPN derivatives as SULT1E1substrates.After molecular docking and experimental validation,an isoform-specific fluorescent substrate(HPN10) for SULT1E1 was successfully engineered.HPN10 demonstrated exceptional isoform-specificity,ultra-high sensitivity,and favorable signal-to-noise ratio(212).HPN10 excelled in the precise sensing of SULT1E1 activities in complex biological matrices,including cellular specimens and liver preparations.HPN10 immensely facilitated the discovery and characterization of SULT1E1 inhibitors,while tetrabromobisphenol A(TBBPA,half inhibitory concentration(IC_(50)) = 31.5 ± 3.4 nmol/L) was identified as a potent SULT1E1 inhibitor that could strongly block SULT1E1 activities in living cells.Collectively,this work presents a practical and efficient strategy for the rational engineering of isoform-specific fluorescent substrates for target conjugative enzyme(s),while HPN10 emerges as a reliable SULT1E1-activatable tool for functional sensing and drug discovery.
