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Fusion of binding domains to thermobifida cellulosilytica cutinase to tune sorption characteristics and enhancing PET hydrolysisRibitsch, Doris ...A cutinase from Thermomyces cellullosylitica (Thc_Cut1), hydrolyzing the synthetic polymer polyethylene terephthalate (PET), was fused with two different binding modules to improve sorption and ... thereby hydrolysis. The binding modules were from cellobiohydrolase I from Hypocrea jecorina (CBM) and from a polyhydroxyalkanoate depolymerase from Alcaligenes faecalis (PBM). Although both binding modules have a hydrophobic nature, it was possible to express the proteins in E. coli. Both fusion enzymes and the native one had comparable kcat values in the range of 311 to 342 s-1 on pNP-butyrate, while the catalytic efficiencies kcat/Km decreased from 0.41 s-1/ M (native enzyme) to 0.21 and 0.33 s-1/ M for Thc_Cut1+PBM and Thc_Cut1+CBM, respectively. The fusion enzymes were active both on the insoluble PET model substrate bis(benzoyloxyethyl) terephthalate (3PET) and on PET although the hydrolysis pattern was differed when compared to Thc_Cut1. Enhanced adsorptionof the fusion enzymes was visible by chemiluminescence after incubation with a 6xHisTag specific horseradish peroxidase (HRP) labeled probe. Increased adsorption to PET by the fusion enzymes was confirmed with Quarz Crystal Microbalance (QCM-D) analysis and indeed resulted in enhanced hydrolysis activity (3.8* for Thc_Cut1+CBM) on PET, as quantified, based on released mono/oligomers.Source: Biomacromolecules. - ISSN 1525-7797 (14, 6, 2013, str. 1769-1776)Type of material - article, component partPublish date - 2013Language - englishCOBISS.SI-ID - 17034518
Author
Ribitsch, Doris |
Orcal Yebra, Antonio |
Zitzenbacher, Sabine |
Ying, Wu, polimeri |
Nowitsch, Susanne |
Steinkellner, Georg |
Greimel, Katrin J. |
Doliška, Aleš |
Oberdorfer, Gustav |
Gruber, Christian C. |
Gruber, Karl, Polimeri |
Schwab, Helmut |
Stana-Kleinschek, Karin |
Herrero Acero, Enrique |
Gübitz, Georg M.
Topics
enzymes |
polymer polyethylene terephthalate |
PET |
hydrolysis |
hydrolysis
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Ribitsch, Doris | ![]() |
Orcal Yebra, Antonio | ![]() |
Zitzenbacher, Sabine | ![]() |
Ying, Wu, polimeri | ![]() |
Nowitsch, Susanne | ![]() |
Steinkellner, Georg | ![]() |
Greimel, Katrin J. | ![]() |
Doliška, Aleš | 17577 |
Oberdorfer, Gustav | ![]() |
Gruber, Christian C. | ![]() |
Gruber, Karl, Polimeri | ![]() |
Schwab, Helmut | ![]() |
Stana-Kleinschek, Karin | 07814 |
Herrero Acero, Enrique | ![]() |
Gübitz, Georg M. | ![]() |
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