Characterization of novel cholesterol esterase from Trichoderma sp. AS59 with high ability to synthesize steryl esters.
A novel cholesterol esterase with there and throughout to synthesisze steryl ester was obtained from the culture filtrate of a fungal strain Trichoderma sp. AS59 isolated from soil. The extracellular enzyme was a monomeric protein with a molecular mass of approximately 58 kDa and an isoelectric point of 4.3. The optimal temperature was between 35 degrees C and 40 degrees C, and the optimal pH was 7.0. The enzyme retained 75% of the initial activity after 18 h of incubation at 30 degrees C in the pH range of 3.5-7.5. Its relative hydrolytic activities on fatty acid cholesteryl esters were in the following order: butyrate (121%), linoleate (100%), caprylate (79%), myristate (42%), palmitate (38%), caproate (37%), and laurate (35%). Unlike mammalian pancreatic cholesterol esterase that is activated by primary cholates on hydrolysis of long-chain fatty acid cholesteryl esters, the enzyme from Trichoderma sp. AS59 displayed its basal activity and was not affected by cholate up to a concentration of 5 mM. At higher cholate concentrations the activity gradually decreased, but reincreased at about 40 mM to reach more than twice the basal activity at 100 mM. The enzyme exhibited a broad substrate specificity, being capable of hydrolyzing various fatty acid esters of not only cholesterol, but also methanol, glycerol, and p-nitrophenol. When incubated with a mixture of cholesterol and oleic acid of equal amounts, the enzyme achieved stoichiometrical esterification in 5 h, indicating its potential utility in food additives and liquid crystal devices.
Maeda
Atsushi
A
Faculty of Engineering, Tokushima Bunri University, 1314-1 Shido, Sanuki 769-2193, Japan. maeda@fe.bunri-u.ac.jp
Mizuno
Takayuki
T
Bunya
Masanori
M
Sugihara
Shigeo
S
Nakayama
Daisuke
D
Tsunasawa
Susumu
S
Hirota
Yoshinori
Y
Sugihara
Akio
A
D020355:Cholates / Q000737:chemistry; D002784:Cholesterol / Q000737:chemistry*; D002788:Cholesterol Esters / Q000138:chemical synthesis* / Q000737:chemistry; D005656:Fungal Proteins / Q000737:chemistry*; D019301:Oleic Acid / Q000737:chemistry*; D002787:Sterol Esterase / Q000737:chemistry*; D013379:Substrate Specificity / Q000502:physiology; D014242:Trichoderma / Q000201:enzymology*
D020355:Cholates; D002788:Cholesterol Esters; D005656:Fungal Proteins; D019301:Oleic Acid; D002784:Cholesterol; D002787:Sterol Esterase