Human cytosolic sulfotransferase 2 A1(SULT2A1) is an important phase II metabolic enzyme. The detection of SULT2A1 is helpful for the functional characterization of SULT2A1 and diagnosis of its related diseases. Howev...Human cytosolic sulfotransferase 2 A1(SULT2A1) is an important phase II metabolic enzyme. The detection of SULT2A1 is helpful for the functional characterization of SULT2A1 and diagnosis of its related diseases. However, due to the overlapping substrate specificity among members of the sulfotransferase family, it is difficult to develop a probe substrate for selective detection of SULT2A1. In the present study, through characterization of the sulfation of series of bufadienolides, arenobufagin(AB) was proved as a potential probe substrate for SULT2A1 with high sensitivity and specificity. Subsequently, the sulfation of AB was characterized by experimental and molecular docking studies. The sulfate-conjugated metabolite was identified as AB-3-sulfate.The sulfation of AB displayed a high selectivity for SULT2A1 which was confirmed by in vitro reaction phenotyping assays. The sulfation of AB by human liver cytosols and recombinant SULT2A1 both obeyed Michaelis-Menten kinetics, with similar kinetic parameters. Molecular docking was performed to understand the interaction between AB and SULT2A1, in which the lack of interaction with Met-137 and Tyr-238 of SULT2A1 made it possible to eliminate substrate inhibition of AB sulfation. Finally, the probe was successfully used to determine the activity of SULT2A1 and its isoenzymes in tissue preparations of human and laboratory animals.展开更多
Microemulsions are promising drug delivery systems for the oral administration of poorly watersoluble drugs. However, the evolution of microemulsions in the gastrointestinal tract is still poorly characterized,especia...Microemulsions are promising drug delivery systems for the oral administration of poorly watersoluble drugs. However, the evolution of microemulsions in the gastrointestinal tract is still poorly characterized,especially the structural change of microemulsions under the effect of lipase and mucus. To better understand the fate of microemulsions in the gastrointestinal tract, we applied small-angle X-ray scattering(SAXS) and fluorescence resonance energy transfer(FRET) to monitor the structural change of microemulsions under the effect of lipolysis and mucus. First, the effect of lipolysis on microemulsions was studied by SAXS, which found the generation of liquid crystalline phases. Meanwhile, FRET spectra indicated micelles with smaller particle sizes were generated during lipolysis, which could be affected by CaCl_2, bile salts and lecithin. Then, the effect of mucus on the structural change of lipolysed microemulsions was studied. The results of SAXS and FRET indicated that the liquid crystalline phases disappeared, and more micelles were generated. In summary, we studied the structural change of microemulsions in simulated gastrointestinal conditions by SAXS and FRET, and successfully monitored the appearance and disappearance of the liquid crystalline phases and micelles.展开更多
基金supported by the National Natural Science Foundation of China (22373017, 22303085, and 21973013)the National Key R&D Program of China (2022YFA1503102)+2 种基金the National Natural Science Foundation of Fujian Province, China (2020J02025)Zhejiang Provincial Natural Science Foundation of China (LQ24B030014)the “Chuying Program” for the Top Young Talents of Fujian Province。
基金the NSFC (81503152,81622047,81473334 and 81503201),distinguished professor of Liaoning Provincesponsored by Liaoning BaiQianWan Talents Program Innovationthe National Key Research and Development Program of China (2017YFC1700200) for financial support
文摘Human cytosolic sulfotransferase 2 A1(SULT2A1) is an important phase II metabolic enzyme. The detection of SULT2A1 is helpful for the functional characterization of SULT2A1 and diagnosis of its related diseases. However, due to the overlapping substrate specificity among members of the sulfotransferase family, it is difficult to develop a probe substrate for selective detection of SULT2A1. In the present study, through characterization of the sulfation of series of bufadienolides, arenobufagin(AB) was proved as a potential probe substrate for SULT2A1 with high sensitivity and specificity. Subsequently, the sulfation of AB was characterized by experimental and molecular docking studies. The sulfate-conjugated metabolite was identified as AB-3-sulfate.The sulfation of AB displayed a high selectivity for SULT2A1 which was confirmed by in vitro reaction phenotyping assays. The sulfation of AB by human liver cytosols and recombinant SULT2A1 both obeyed Michaelis-Menten kinetics, with similar kinetic parameters. Molecular docking was performed to understand the interaction between AB and SULT2A1, in which the lack of interaction with Met-137 and Tyr-238 of SULT2A1 made it possible to eliminate substrate inhibition of AB sulfation. Finally, the probe was successfully used to determine the activity of SULT2A1 and its isoenzymes in tissue preparations of human and laboratory animals.
基金supported by the National Natural Science Foundation of China (Grant No. 81703606)the Educational Committee Foundation of Liaoning Province (Grant No. L2016026)the Fundamental Research Funds for the Central Universities (Grant no. wd01185)
文摘Microemulsions are promising drug delivery systems for the oral administration of poorly watersoluble drugs. However, the evolution of microemulsions in the gastrointestinal tract is still poorly characterized,especially the structural change of microemulsions under the effect of lipase and mucus. To better understand the fate of microemulsions in the gastrointestinal tract, we applied small-angle X-ray scattering(SAXS) and fluorescence resonance energy transfer(FRET) to monitor the structural change of microemulsions under the effect of lipolysis and mucus. First, the effect of lipolysis on microemulsions was studied by SAXS, which found the generation of liquid crystalline phases. Meanwhile, FRET spectra indicated micelles with smaller particle sizes were generated during lipolysis, which could be affected by CaCl_2, bile salts and lecithin. Then, the effect of mucus on the structural change of lipolysed microemulsions was studied. The results of SAXS and FRET indicated that the liquid crystalline phases disappeared, and more micelles were generated. In summary, we studied the structural change of microemulsions in simulated gastrointestinal conditions by SAXS and FRET, and successfully monitored the appearance and disappearance of the liquid crystalline phases and micelles.
