Improved reactivation of immobilized-stabilized lipase from Thermomyces lanuginosus by its coating with highly hydrophilic polymers.
Immobilized-stabilized aminated lipase from Thermomyces lanuginosus (TLL-A) is not easily reactivated after inactivation by incubation in the presence of organic solvents or chaotropic reagents. To improve the recovered activity of this biocatalyst, immobilized TLL-A has been submitted to different modifications. The best results were obtained when the enzyme was coated with a very hydrophilic and inert polymer: dextran modified with glycine (Dx-Gly). This modification did not reduce enzymatic activity while it increased the stability of this already very stable preparation, in thermal and organic solvent induced inactivation (by a 4-fold factor). Simple incubation in aqueous medium at pH 7 and 25 degrees C permitted to fully recover the activity of the immobilized and modified TLL-A enzyme inactivated by incubation in organic solvents or saturated guanidine during 3 cycles, while the non-modified enzyme only recover some activity. When the inactivation was caused by exposition at high temperatures, the reactivation was higher using the modified biocatalyst, but was far for complete (40% after 3 inactivation-reactivation cycles). The determination of the TLL-A activity in the presence of detergents (that helps the opening of active site of the lipase) allowed, in this case, to significantly improve the results, now near to 90% of the initial activity was recovered (using the non-modified enzyme the recovered activity was around 60%). This very hydrophilic and inert polymer, coating the enzyme surface, seems to help the correct positioning of the hydrophilic and hydrophobic groups of the enzyme, and that way improve both the stability and possibility of reactivation of the enzyme.
Rodrigues
Rafael C
RC
Departamento de Biocatálisis, Instituto de Catálisis-CSIC, Campus UAM, Cantoblanco, Madrid, Spain.
Bolivar
Juan M
JM
Volpato
Giandra
G
Filice
Marco
M
Godoy
Cesar
C
Fernandez-Lafuente
Roberto
R
Guisan
Jose M
JM
D000588:Amines / Q000378:metabolism; D001203:Ascomycota / Q000201:enzymology*; D003124:Colorimetry; D004789:Enzyme Activation / Q000187:drug effects; D004795:Enzyme Stability / Q000187:drug effects; D004800:Enzymes, Immobilized / Q000378:metabolism*; D006038:Glyoxylates / Q000378:metabolism; D019791:Guanidine / Q000494:pharmacology; D006358:Hot Temperature; D008049:Lipase / Q000378:metabolism*; D011108:Polymers / Q000494:pharmacology*; D012685:Sepharose / Q000378:metabolism; D012997:Solvents / Q000494:pharmacology; D013997:Time Factors; D017186:Titrimetry
D000588:Amines; D004800:Enzymes, Immobilized; D006038:Glyoxylates; D011108:Polymers; D012997:Solvents; C070399:glyoxyl agarose; D012685:Sepharose; D008049:Lipase; D019791:Guanidine