title: Biomimetic Approach to Enhance Enzymatic Hydrolysis of the Synthetic Polyester Poly(1,4-butylene adipate): Fusing Binding Modules to Esterases.
abstract: Mimicking a concept of nature for the hydrolysis of biopolymers, the Thermobifida cellulosilytica cutinase 1 (Thc_Cut1) was fused to a polymer binding module (PBM) to enhance the hydrolysis of the polyester poly(1,4-butylene adipate) (PBA). Namely, the binding module of a polyhydroxyalkanoate depolymerase from Alcaligenes faecalis (Thc_Cut1_PBM) was attached to the cutinase via two different linker sequences varying in length. In order to investigate the adsorption behavior, catalytically inactive mutants both of Thc_Cut1 and Thc_Cut1_PBM were successfully constructed by site-directed mutagenesis of serine 131 to alanine. Quartz crystal microbalance with dissipation monitoring (QCM-D) analysis revealed that the initial mass increase during enzyme adsorption was larger for the inactive enzymes linked with the PBM as compared to the enzyme without the PBM. The hydrolysis rates of PBA were significantly enhanced when incubated with the active, engineered Thc_Cut1_PBM as compared to the native Thc_Cut1. Thc_Cut1_PBM completely hydrolyzed PBA thin films on QCM-D sensors within approximately 40 min, whereas twice as much time was required for the complete hydrolysis by the native Thc_Cut1.
authors.affiliation: Austrian Centre of Industrial Biotechnology, ACIB , Konrad Lorenz Strasse 20, 3430 Tulln, Austria.
authors.forename: Veronika
authors.initials: V
authors.lastname: Perz
authors.affiliation: Department of Environmental Systems Science, Institute of Biogeochemistry and Pollutant Dynamics, Swiss Federal Institute of Technology Zurich (ETHZ) , Universitätstrasse 16, 8092 Zurich, Switzerland.
authors.forename: Michael Thomas
authors.initials: MT
authors.lastname: Zumstein
authors.affiliation: Department of Environmental Systems Science, Institute of Biogeochemistry and Pollutant Dynamics, Swiss Federal Institute of Technology Zurich (ETHZ) , Universitätstrasse 16, 8092 Zurich, Switzerland.
authors.forename: Michael
authors.initials: M
authors.lastname: Sander
authors.affiliation: Austrian Centre of Industrial Biotechnology, ACIB , Petersgasse 14/V, 8010 Graz, Austria.
authors.forename: Sabine
authors.initials: S
authors.lastname: Zitzenbacher
authors.affiliation: Austrian Centre of Industrial Biotechnology, ACIB , Konrad Lorenz Strasse 20, 3430 Tulln, Austria.|Institute of Environmental Biotechnology, University of Natural Resources and Life Sciences, Vienna , Konrad Lorenz Strasse 20, 3430 Tulln, Austria.
authors.forename: Doris
authors.initials: D
authors.lastname: Ribitsch
authors.affiliation: Austrian Centre of Industrial Biotechnology, ACIB , Konrad Lorenz Strasse 20, 3430 Tulln, Austria.|Institute of Environmental Biotechnology, University of Natural Resources and Life Sciences, Vienna , Konrad Lorenz Strasse 20, 3430 Tulln, Austria.
authors.forename: Georg M
authors.initials: GM
authors.lastname: Guebitz
mesh_terms: D039903:Actinobacteria / Q000737:chemistry / Q000201:enzymology; D000409:Alanine / Q000737:chemistry / Q000235:genetics; D042481:Alcaligenes faecalis / Q000737:chemistry / Q000201:enzymology; D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D032701:Biomimetics / Q000379:methods*; D002072:Butylene Glycols / Q000737:chemistry* / Q000378:metabolism; D002265:Carboxylic Ester Hydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D006868:Hydrolysis; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D011108:Polymers / Q000737:chemistry* / Q000378:metabolism; D015202:Protein Engineering; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012694:Serine / Q000737:chemistry / Q000235:genetics; D013329:Structure-Activity Relationship
chemical_list: D001426:Bacterial Proteins; D002072:Butylene Glycols; D011108:Polymers; D011994:Recombinant Proteins; D012694:Serine; C009204:butylene glycol adipic acid polyester; D002265:Carboxylic Ester Hydrolases; C019952:cutinase; C077226:poly(3-hydroxyalkanoic acid) depolymerase; D000409:Alanine