Biomaterial Database

Affinity purification of cytosolic epoxide hydrolase using derivatized epoxy-activated Sepharose gels. 1988

R N Wixtrom, and M H Silva, and B D Hammock

Department of Entomology, University of California, Davis 95616.

Improved affinity chromatography procedures for the purification of cytosolic epoxide hydrolase are described. An earlier affinity purification method using immobilized 7-methoxycitronellyl thiol (MCT) sporadically produced final enzyme preparations containing major impurities. To eliminate these impurities, we tested alternate ligands, spacer arms, and ligand concentrations. A series of alkyl and aryl thiols coupled to epoxy-activated Sepharose were found to exhibit markedly different binding characteristics as compared with commercially available alkyl- and aryl-Sepharose gels. Using one of these new matrices, benzylthio-Sepharose, cytosolic epoxide hydrolase from mouse liver was purified over 100-fold, appeared homogeneous by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and was obtained with 60-90% recovery of enzyme activity. The impurities previously observed with the MCT-Sepharose procedure were reduced or eliminated by using an MCT ligand concentration of 5 microequivalents per gram or less. MCT-Sepharose and benzylthio-Sepharose provide rapid and convenient one-step procedures for obtaining purified cytosolic epoxide hydrolase from numerous species and tissues.

UI	MeSH Term	Description	Entries
D008099	Liver	A large lobed glandular organ in the abdomen of vertebrates that is responsible for detoxification, metabolism, synthesis and storage of various substances.	Livers
D008253	Macaca mulatta	A species of the genus MACACA inhabiting India, China, and other parts of Asia. The species is used extensively in biomedical research and adapts very well to living with humans.	Chinese Rhesus Macaques,Macaca mulatta lasiota,Monkey, Rhesus,Rhesus Monkey,Rhesus Macaque,Chinese Rhesus Macaque,Macaca mulatta lasiotas,Macaque, Rhesus,Rhesus Macaque, Chinese,Rhesus Macaques,Rhesus Macaques, Chinese,Rhesus Monkeys
D008297	Male		Males
D002846	Chromatography, Affinity	A chromatographic technique that utilizes the ability of biological molecules, often ANTIBODIES, to bind to certain ligands specifically and reversibly. It is used in protein biochemistry. (McGraw-Hill Dictionary of Scientific and Technical Terms, 4th ed)	Chromatography, Bioaffinity,Immunochromatography,Affinity Chromatography,Bioaffinity Chromatography
D003600	Cytosol	Intracellular fluid from the cytoplasm after removal of ORGANELLES and other insoluble cytoplasmic components.	Cytosols
D003956	Dialysis	A process of selective diffusion through a membrane. It is usually used to separate low-molecular-weight solutes which diffuse through the membrane from the colloidal and high-molecular-weight solutes which do not. (McGraw-Hill Dictionary of Scientific and Technical Terms, 4th ed)	Dialyses
D004591	Electrophoresis, Polyacrylamide Gel	Electrophoresis in which a polyacrylamide gel is used as the diffusion medium.	Polyacrylamide Gel Electrophoresis,SDS-PAGE,Sodium Dodecyl Sulfate-PAGE,Gel Electrophoresis, Polyacrylamide,SDS PAGE,Sodium Dodecyl Sulfate PAGE,Sodium Dodecyl Sulfate-PAGEs
D004795	Enzyme Stability	The extent to which an enzyme retains its structural conformation or its activity when subjected to storage, isolation, and purification or various other physical or chemical manipulations, including proteolytic enzymes and heat.	Enzyme Stabilities,Stabilities, Enzyme,Stability, Enzyme
D004851	Epoxide Hydrolases	Enzymes that catalyze reversibly the formation of an epoxide or arene oxide from a glycol or aromatic diol, respectively.	Epoxide Hydrase,Epoxide Hydrases,Epoxide Hydratase,Epoxide Hydratases,Epoxide Hydrolase,9,10-Epoxypalmitic Acid Hydrase,Microsomal Epoxide Hydrolase,Styrene Epoxide Hydrolase,9,10 Epoxypalmitic Acid Hydrase,Acid Hydrase, 9,10-Epoxypalmitic,Epoxide Hydrolase, Microsomal,Epoxide Hydrolase, Styrene,Hydrase, 9,10-Epoxypalmitic Acid,Hydrase, Epoxide,Hydrases, Epoxide,Hydratase, Epoxide,Hydratases, Epoxide,Hydrolase, Epoxide,Hydrolase, Microsomal Epoxide,Hydrolase, Styrene Epoxide,Hydrolases, Epoxide
D004852	Epoxy Compounds	Organic compounds that include a cyclic ether with three ring atoms in their structure. They are commonly used as precursors for POLYMERS such as EPOXY RESINS.	Epoxide,Epoxides,Epoxy Compound,Oxiranes,Compound, Epoxy,Compounds, Epoxy