A comparison of the kinetic properties of free and immobilized Aspergillus oryzae β-galactosidase

被引:39
作者
Freitas, Fernanda F. [1 ]
Marquez, Libia D. S. [1 ]
Ribeiro, Gustavo P. [1 ]
Brandao, Gabriela C. [1 ]
Cardoso, Vicelma L. [1 ]
Ribeiro, Eloizio J. [1 ]
机构
[1] Univ Fed Uberlandia, Fac Chem Engn, BR-38400902 Uberlandia, MG, Brazil
关键词
Immobilized enzymes; beta-galactosidase; Alginate; Glucose; Kinetic parameters; Lactose; GLUTARALDEHYDE CROSS-LINKING; LACTOSE HYDROLYSIS; PRODUCT INHIBITION; WHEY PERMEATE; SUPPORTS; CHITOSAN; STABILITY; BEADS;
D O I
10.1016/j.bej.2011.08.011
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
The objective of this work was to compare the properties of free and immobilized beta-galactosiclase (Aspergillus oryzae), entrapped in alginate-gelatin beads and cross-linked with glutaraldehyde. The free and immobilized forms of the enzyme showed no decrease in enzyme activity when incubated in buffer solutions in pH ranges of 4.5-7.0. The kinetics of lactose hydrolysis by the free and immobilized enzymes were studied at maximum substrate concentrations of 90 g/L and 140 g/L, respectively, a temperature of 35 degrees C and a pH of 4.5. The Michaelis-Menten model with competitive inhibition by galactose fit the experimental results for both forms. The K-m and V-m values of the free enzyme were 52.13 +/- 2.8 mM and 2.56 +/- 0.3 g(glucose)/L min mg(enzyme), respectively, and were 60.30 +/- 3.3 mM and 1032.07 +/- 51.6 g(lactose)/min m(catalyst)(3), respectively, for the immobilized form. The maximum enzymatic activity of the soluble form of eta-galactosidase was obtained at pH 4.5 and 55 degrees C. Alternatively, the immobilized form was most active at pH 5.0 at 60 degrees C. The free and immobilized enzymes presented activation energies of 6.90 +/- 0.5 kcal/mol and 7.7 +/- 0.7 kcal/mol, respectively, which suggested that the immobilized enzyme possessed a lower resistance to substrate transfer. (C) 2011 Elsevier B.V. All rights reserved.
引用
收藏
页码:33 / 38
页数:6
相关论文
共 47 条
[1]   Scientific Opinion on lactose thresholds in lactose intolerance and galactosaemia [J].
Agostoni, Carlo ;
Bresson, Jean-Louis ;
Fairweather-Tait, Susan ;
Flynn, Albert ;
Golly, Ines ;
Korhonen, Harmu ;
Lagiou, Pagona ;
Lovik, Martinus ;
Marchelli, Rosangela ;
Martin, Ambroise ;
Moseley, Bevan ;
Neuhauser-Berthold, Monika ;
Przyrembel, Hildegard ;
Salminen, Seppo ;
Sanz, Yolanda ;
Strain, Sean ;
Strobel, Stephan ;
Tetens, Inge ;
Tome, Daniel ;
van Loveren, Hendrik ;
Verhagen, Hans .
EFSA JOURNAL, 2010, 8 (09)
[2]   Effects of internal mass transfer and product inhibition on a simulated immobilized enzyme-catalyzed reactor for lactose hydrolysis [J].
Al-Muftah, AE ;
Abu-Reesh, IM .
BIOCHEMICAL ENGINEERING JOURNAL, 2005, 23 (02) :139-153
[3]  
Ansari S. A., 2010, J MOL CATAL-B ENZYM, V63, P93
[4]   A new method for immobilization of beta-galactosidase and its utilization in a plug flow reactor [J].
Ates, S ;
Mehmetoglu, U .
PROCESS BIOCHEMISTRY, 1997, 32 (05) :433-436
[5]   Glucose oxidase - An overview [J].
Bankar, Sandip B. ;
Bule, Mahesh V. ;
Singhal, Rekha S. ;
Ananthanarayan, Laxmi .
BIOTECHNOLOGY ADVANCES, 2009, 27 (04) :489-501
[6]   Different mechanisms of protein immobilization on glutaraldehyde activated supports:: Effect of support activation and immobilization conditions [J].
Betancor, Lorena ;
Lopez-Gallego, Fernando ;
Hidalgo, Aurelio ;
Alonso-Morales, Noelia ;
Dellamora-Ortiz Cesar Mateo, Gisela ;
Fernandez-Lafuente, Roberto ;
Guisan, Jose M. .
ENZYME AND MICROBIAL TECHNOLOGY, 2006, 39 (04) :877-882
[7]  
Bulchholz K., 2005, BIOCATALYSTS ENZYME
[8]   Immobilised enzymes: science or art? [J].
Cao, LQ .
CURRENT OPINION IN CHEMICAL BIOLOGY, 2005, 9 (02) :217-226
[9]  
Chibata I., 1978, IMMOBILISED ENZYME R
[10]   Chemically surface modified gel (CSMG): An excellent enzyme-immobilization matrix for industrial processes [J].
David, Allan E. ;
Wang, Nam Sun ;
Yang, Victor C. ;
Yang, Arthur J. .
JOURNAL OF BIOTECHNOLOGY, 2006, 125 (03) :395-407