Purification and characterization of a feruloyl esterase from Fusarium oxysporum catalyzing esterification of phenolic acids in ternary water-organic solvent mixtures.
An extracellular feruloyl esterase (FAE-II) from the culture filtrates of Fusarium oxysporum F3 was purified to homogeneity by SP-Sepharose, t-butyl-HIC and Sephacryl S-200 column chromatography. The protein corresponded to molecular mass and pI values of 27 kDa and 9.9, respectively. The enzyme was optimally active at pH 7 and 45 degrees C. The purified esterase was fully stable at pH 7.0-9.0 and temperature up to 45 degrees C after 1 h incubation. Determination of k(cat)/K(m) revealed that the enzyme hydrolysed methyl sinapinate 6, 21 and 40 times more efficiently than methyl ferulate, methyl coumarate and methyl caffeate, respectively. The enzyme was active on substrates containing ferulic acid ester linked to the C-5 but inactive to the C-2 positions of arabinofuranose such as 4-nitrophenyl 5-O-trans-feruloyl-alpha-L-arabinofuranoside and 4-nitrophenyl 2-O-trans-feruloyl-alpha-L-arabinofuranoside. In the presence of Sporotrichum thermophile xylanase, there was a significant release of ferulic acid from destarched wheat bran by FAE-II, indicating a synergistic interaction between FAE-II and S. thermophile xylanase. FAE-II by itself could release only little ferulic acid from destarched wheat bran. The potential of FAE-II for the synthesis of various phenolic acid esters was tested using as a reaction system a surfactantless microemulsion formed in ternary mixture consisting of n-hexane, 1-propanol and water.
Topakas
E
E
Chemical Engineering Department, National Technical University of Athens, 5 Iroon Polytechniou Str, Zografou Campus, 15700 Athens, Greece.
Stamatis
H
H
Biely
P
P
Kekos
D
D
Macris
B J
BJ
Christakopoulos
P
P
D001089:Arabinose / Q000031:analogs & derivatives* / Q000737:chemistry; D002265:Carboxylic Ester Hydrolases / Q000096:biosynthesis / Q000737:chemistry* / Q000145:classification / Q000302:isolation & purification*; D002384:Catalysis; D002847:Chromatography, Agarose; D002934:Cinnamates / Q000737:chemistry; D003067:Coenzymes / Q000737:chemistry; D003373:Coumaric Acids / Q000138:chemical synthesis* / Q000737:chemistry; D004655:Emulsions / Q000737:chemistry; D004789:Enzyme Activation; D004795:Enzyme Stability; D004951:Esterification; D005670:Fusarium / Q000737:chemistry* / Q000145:classification / Q000201:enzymology; D006863:Hydrogen-Ion Concentration; D062385:Hydroxybenzoates / Q000737:chemistry*; D008970:Molecular Weight; D009930:Organic Chemicals / Q000737:chemistry; D012997:Solvents / Q000737:chemistry; D013045:Species Specificity; D013175:Sporothrix / Q000737:chemistry* / Q000145:classification / Q000201:enzymology; D013379:Substrate Specificity; D013696:Temperature; D014867:Water / Q000737:chemistry; D014995:Xylosidases / Q000737:chemistry* / Q000378:metabolism
D002934:Cinnamates; D003067:Coenzymes; D003373:Coumaric Acids; D004655:Emulsions; D062385:Hydroxybenzoates; D009930:Organic Chemicals; D012997:Solvents; C431714:arabinofuranose; D014867:Water; C006605:antithiamine factor; C004999:ferulic acid; D001089:Arabinose; D002265:Carboxylic Ester Hydrolases; C070196:feruloyl esterase; D014995:Xylosidases; C017616:phenolic acid