title: Kinetic properties and solubilization of microsomal cholesterol ester hydrolase from rat liver.
abstract: Some kinetic properties of the microsomal cholesterol ester hydrolase (CEH) have been examined in rat liver. The reaction was linear with time up to 60 min and with enzyme concentration up to 0.3 mg/mL, and a pH optimum of 6.7 for enzyme activity was observed. Cholesterol esterase exhibited the following apparent kinetic constants: Km, 68.88 microM and Vmax, 33 Units/mg protein. Cholesteryl palmitate was hydrolyzed to a much greater extent than cholesteryl oleate by the enzyme. Product inhibition with cholesterol and palmitic acid was not apparent; however, oleic acid added to the system reduced markedly microsomal CEH activity. The present paper also reports the solubilization of cholesteryl palmitate hydrolase from the microsomal fraction by pretreating it with Triton X-100, sodium deoxycholate, and sodium dodecylsulfate. All ionic and non-ionic detergents tested are capable of making the enzyme soluble, and maximal effects were found at higher concentrations of detergents although the esterase activity was strongly inhibited. Triton X-100 was found to be more effective than sodium deoxycholate and sodium dodecylsulfate in enzyme and protein solubilization. When the direct effects of detergents on CEH activity were studied, progressive concentration-dependent inhibitions were observed.
authors.affiliation: Department of Physiology and Biochemistry, University of the Basque Country Medical School, Bilbao, Spain.
authors.forename: J M
authors.initials: JM
authors.lastname: Gandarias
authors.forename: M
authors.initials: M
authors.lastname: Lacort
authors.forename: M J
authors.initials: MJ
authors.lastname: Martinez
authors.forename: C J
authors.initials: CJ
authors.lastname: Fernandez
authors.forename: B
authors.initials: B
authors.lastname: Ochoa
mesh_terms: D000818:Animals; D002265:Carboxylic Ester Hydrolases / Q000378:metabolism*; D002788:Cholesterol Esters / Q000378:metabolism*; D005260:Female; D006868:Hydrolysis; D007700:Kinetics; D008862:Microsomes, Liver / Q000201:enzymology*; D051381:Rats; D011919:Rats, Inbred Strains; D012995:Solubility; D002787:Sterol Esterase / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D002788:Cholesterol Esters; D002265:Carboxylic Ester Hydrolases; D002787:Sterol Esterase