title: Application of Frontal Affinity Chromatography to Study the Biomolecular Interactions with Trypsin.
abstract: Trypsin is a serine protease that has been proposed as a potential therapeutic target for metabolic disorders and malignancy diseases, thus the identification of biomolecular interactions of compounds to trypsin could be of great therapeutic importance. In this study, trypsin was immobilized on a monolithic silica capillary column via sol-gel. The binding properties of four small molecules (daidzin, genistin, matrine and oxymatrine) to trypsin were examined using the trypsin affinity columns by frontal analysis. The results indicate that the matrine (dissociation constant, Kd = 7.904 μM) has stronger interaction with trypsin than the oxymatrine (Kd = 8.204 μM), whereas daidzin and genistin were nearly have no affinity with trypsin. The results demonstrated that the frontal affinity chromatography can be used for the direct determination of protein-protease inhibitor binding interactions and have several significant advantages, including easy fabricating, reproducible, minimal technological requirements and potential to become a reliable alternative for quantitative studies of biomolecular interactions.
authors.affiliation: Department of Pharmaceutical Science, Zhejiang University of Technology, Zhejiang 310014, PR China.
authors.forename: YuanYuan
authors.initials: Y
authors.lastname: Hu
authors.affiliation: Department of Pharmaceutical Science, Zhejiang University of Technology, Zhejiang 310014, PR China pharmo@163.com.
authors.forename: Junqing
authors.initials: J
authors.lastname: Qian
authors.affiliation: Department of Pharmaceutical Science, Zhejiang University of Technology, Zhejiang 310014, PR China.
authors.forename: Hui
authors.initials: H
authors.lastname: Guo
authors.affiliation: Department of Pharmaceutical Science, Zhejiang University of Technology, Zhejiang 310014, PR China.
authors.forename: ShengLan
authors.initials: S
authors.lastname: Jiang
authors.affiliation: Department of Pharmaceutical Science, Zhejiang University of Technology, Zhejiang 310014, PR China.
authors.forename: Zheng
authors.initials: Z
authors.lastname: Zhang
mesh_terms: D000470:Alkaloids; D002846:Chromatography, Affinity / Q000379:methods*; D004800:Enzymes, Immobilized / Q000737:chemistry / Q000378:metabolism*; D007529:Isoflavones; D007700:Kinetics; D008956:Models, Chemical; D011807:Quinolizines; D012822:Silicon Dioxide; D014357:Trypsin / Q000737:chemistry / Q000378:metabolism*; D014361:Trypsin Inhibitors / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism*; D000093842:Matrines
chemical_list: D000470:Alkaloids; D004800:Enzymes, Immobilized; D007529:Isoflavones; D011807:Quinolizines; D014361:Trypsin Inhibitors; C040641:genistin; C013908:daidzin; D012822:Silicon Dioxide; C037573:oxymatrine; D014357:Trypsin; D000093842:Matrines