Perz V

References (8)

Title : Enzymatic Hydrolysis of Polyester Thin Films at the Nanoscale: Effects of Polyester Structure and Enzyme Active-Site Accessibility - Zumstein_2017_Environ.Sci.Technol_51_7476
Author(s) : Zumstein MT , Rechsteiner D , Roduner N , Perz V , Ribitsch D , Guebitz GM , Kohler HE , McNeill K , Sander M
Ref : Environ Sci Technol , 51 :7476 , 2017
Abstract : Biodegradable polyesters have a large potential to replace persistent polymers in numerous applications and to thereby reduce the accumulation of plastics in the environment. Ester hydrolysis by extracellular carboxylesterases is considered the rate-limiting step in polyester biodegradation. In this work, we systematically investigated the effects of polyester and carboxylesterase structure on the hydrolysis of nanometer-thin polyester films using a quartz-crystal microbalance with dissipation monitoring. Hydrolyzability increased with increasing polyester-chain flexibility as evidenced from differences in the hydrolysis rates and extents of aliphatic polyesters varying in the length of their dicarboxylic acid unit and of poly(butylene adipate-co-terephthalate) (PBAT) polyesters varying in their terephthalate-to-adipate ratio by Rhizopus oryzae lipase and Fusarium solani cutinase. Nanoscale nonuniformities in the PBAT films affected enzymatic hydrolysis and were likely caused by domains with elevated terephthalate contents that impaired enzymatic hydrolysis. Yet, the cutinase completely hydrolyzed all PBAT films, including films with a terephthalate-to-adipate molar ratio of one, under environmentally relevant conditions (pH 6, 20 degrees C). A comparative analysis of the hydrolysis of two model polyesters by eight different carboxylesterases revealed increasing hydrolysis with increasing accessibility of the enzyme active site. Therefore, this work highlights the importance of both polyester and carboxylesterase structure to enzymatic polyester hydrolysis.
ESTHER : Zumstein_2017_Environ.Sci.Technol_51_7476
PubMedSearch : Zumstein_2017_Environ.Sci.Technol_51_7476
PubMedID: 28538100

Title : Discovery of Polyesterases from Moss-Associated Microorganisms - Muller_2017_Appl.Environ.Microbiol_83_e02641
Author(s) : Muller CA , Perz V , Provasnek C , Quartinello F , Guebitz GM , Berg G
Ref : Applied Environmental Microbiology , 83 : , 2017
Abstract : The growing pollution of the environment with plastic debris is a global threat which urgently requires biotechnological solutions. Enzymatic recycling not only prevents pollution but also would allow recovery of valuable building blocks. Therefore, we explored the existence of microbial polyesterases in microbial communities associated with the Sphagnum magellanicum moss, a key species within unexploited bog ecosystems. This resulted in the identification of six novel esterases, which were isolated, cloned, and heterologously expressed in Escherichia coli The esterases were found to hydrolyze the copolyester poly(butylene adipate-co-butylene terephthalate) (PBAT) and the oligomeric model substrate bis[4-(benzoyloxy)butyl] terephthalate (BaBTaBBa). Two promising polyesterase candidates, EstB3 and EstC7, which clustered in family VIII of bacterial lipolytic enzymes, were purified and characterized using the soluble esterase substrate p-nitrophenyl butyrate (K(m) values of 46.5 and 3.4 microM, temperature optima of 48 degreesC and 50 degreesC, and pH optima of 7.0 and 8.5, respectively). In particular, EstC7 showed outstanding activity and a strong preference for hydrolysis of the aromatic ester bond in PBAT. Our study highlights the potential of plant-associated microbiomes from extreme natural ecosystems as a source for novel hydrolytic enzymes hydrolyzing polymeric compounds. IMPORTANCE: In this study, we describe the discovery and analysis of new enzymes from microbial communities associated with plants (moss). The recovered enzymes show the ability to hydrolyze not only common esterase substrates but also the synthetic polyester poly(butylene adipate-co-butylene terephthalate), which is a common material employed in biodegradable plastics. The widespread use of such synthetic polyesters in industry and society requires the development of new sustainable technological solutions for their recycling. The discovered enzymes have the potential to be used as catalysts for selective recovery of valuable building blocks from this material.
ESTHER : Muller_2017_Appl.Environ.Microbiol_83_e02641
PubMedSearch : Muller_2017_Appl.Environ.Microbiol_83_e02641
PubMedID: 27940546
Gene_locus related to this paper: 9bact-a0a1c9t7d3 , 9bact-a0a1c9t7d1

Title : Characterization of a poly(butylene adipate-co-terephthalate)-hydrolyzing lipase from Pelosinus fermentans - Biundo_2016_Appl.Microbiol.Biotechnol_100_1753
Author(s) : Biundo A , Hromic A , Pavkov-Keller T , Gruber K , Quartinello F , Haernvall K , Perz V , Arrell MS , Zinn M , Ribitsch D , Guebitz GM
Ref : Applied Microbiology & Biotechnology , 100 :1753 , 2016
Abstract : Certain alpha/beta hydrolases have the ability to hydrolyze synthetic polyesters. While their partial hydrolysis has a potential for surface functionalization, complete hydrolysis allows recycling of valuable building blocks. Although knowledge about biodegradation of these materials is important regarding their fate in the environment, it is currently limited to aerobic organisms. A lipase from the anaerobic groundwater organism Pelosinus fermentans DSM 17108 (PfL1) was cloned and expressed in Escherichia coli BL21-Gold(DE3) and purified from the cell extract. Biochemical characterization with small substrates showed thermoalkalophilic properties (T opt = 50 degrees C, pHopt = 7.5) and higher activity towards para-nitrophenyl octanoate (12.7 U mg-1) compared to longer and shorter chain lengths (C14 0.7 U mg-1 and C2 4.3 U mg-1, respectively). Crystallization and determination of the 3-D structure displayed the presence of a lid structure and a zinc ion surrounded by an extra domain. These properties classify the enzyme into the I.5 lipase family. PfL1 is able to hydrolyze poly(1,4-butylene adipate-co-terephthalate) (PBAT) polymeric substrates. The hydrolysis of PBAT showed the release of small building blocks as detected by liquid chromatography-mass spectrometry (LC-MS). Protein dynamics seem to be involved with lid opening for the hydrolysis of PBAT by PfL1.
ESTHER : Biundo_2016_Appl.Microbiol.Biotechnol_100_1753
PubMedSearch : Biundo_2016_Appl.Microbiol.Biotechnol_100_1753
PubMedID: 26490551
Gene_locus related to this paper: 9firm-a0a0a0ymq9

Title : Data on synthesis of oligomeric and polymeric poly(butylene adipate-co-butylene terephthalate) model substrates for the investigation of enzymatic hydrolysis - Perz_2016_Data.Brief_7_291
Author(s) : Perz V , Bleymaier K , Sinkel C , Kueper U , Bonnekessel M , Ribitsch D , Guebitz GM
Ref : Data Brief , 7 :291 , 2016
Abstract : The aliphatic-aromatic copolyester poly(butylene adipate-co-butylene terephthalate) (PBAT), also known as ecoflex, contains adipic acid, 1,4-butanediol and terephthalic acid and is proven to be compostable [1], [2], [3]). We describe here data for the synthesis and analysis of poly(butylene adipate-co-butylene terephthalate variants with different adipic acid:terephatalic acid ratios and 6 oligomeric PBAT model substrates. Data for the synthesis of the following oligomeric model substrates are described: mono(4-hydroxybutyl) terephthalate (BTa), bis(4-(hexanoyloxy)butyl) terephthalate (HaBTaBHa), bis(4-(decanoyloxy)butyl) terephthalate (DaBTaBDa), bis(4-(tetradecanoyloxy)butyl) terephthalate (TdaBTaBTda), bis(4-hydroxyhexyl) terephthalate (HTaH) and bis(4-(benzoyloxy)butyl) terephthalate (BaBTaBBa). Polymeric PBAT variants were synthesized with adipic acid:terephatalic acid ratios of 100:0, 90:10, 80:20, 70:30, 60:40 and 50:50. These polymeric and oligomeric substances were used as ecoflex model substrates in enzymatic hydrolysis experiments in the article "Substrate specificities of cutinases on aliphatic-aromatic polyesters and on their model substrates" [4].
ESTHER : Perz_2016_Data.Brief_7_291
PubMedSearch : Perz_2016_Data.Brief_7_291
PubMedID: 26981550

Title : An Esterase from Anaerobic Clostridium hathewayi Can Hydrolyze Aliphatic-Aromatic Polyesters - Perz_2016_Environ.Sci.Technol_50_2899
Author(s) : Perz V , Hromic A , Baumschlager A , Steinkellner G , Pavkov-Keller T , Gruber K , Bleymaier K , Zitzenbacher S , Zankel A , Mayrhofer C , Sinkel C , Kueper U , Schlegel K , Ribitsch D , Guebitz GM
Ref : Environ Sci Technol , 50 :2899 , 2016
Abstract : Recently, a variety of biodegradable polymers have been developed as alternatives to recalcitrant materials. Although many studies on polyester biodegradability have focused on aerobic environments, there is much less known on biodegradation of polyesters in natural and artificial anaerobic habitats. Consequently, the potential of anaerobic biogas sludge to hydrolyze the synthetic compostable polyester PBAT (poly(butylene adipate-co-butylene terephthalate) was evaluated in this study. On the basis of reverse-phase high-performance liquid chromatography (RP-HPLC) analysis, accumulation of terephthalic acid (Ta) was observed in all anaerobic batches within the first 14 days. Thereafter, a decline of Ta was observed, which occurred presumably due to consumption by the microbial population. The esterase Chath_Est1 from the anaerobic risk 1 strain Clostridium hathewayi DSM-13479 was found to hydrolyze PBAT. Detailed characterization of this esterase including elucidation of the crystal structure was performed. The crystal structure indicates that Chath_Est1 belongs to the alpha/beta-hydrolases family. This study gives a clear hint that also micro-organisms in anaerobic habitats can degrade manmade PBAT.
ESTHER : Perz_2016_Environ.Sci.Technol_50_2899
PubMedSearch : Perz_2016_Environ.Sci.Technol_50_2899
PubMedID: 26878094
Gene_locus related to this paper: 9clot-r5t6k9

Title : Substrate specificities of cutinases on aliphatic-aromatic polyesters and on their model substrates - Perz_2016_N.Biotechnol_33_295
Author(s) : Perz V , Bleymaier K , Sinkel C , Kueper U , Bonnekessel M , Ribitsch D , Guebitz GM
Ref : N Biotechnol , 33 :295 , 2016
Abstract : The enzymatic hydrolysis of the biodegradable polyester ecoflex and of a variety of oligomeric and polymeric ecoflex model substrates was investigated. For this purpose, substrate specificities of two enzymes of typical compost inhabitants, namely a fungal cutinase from Humicola insolens (HiC) and a bacterial cutinase from Thermobifida cellulosilytica (Thc_Cut1) were compared. Model substrates were systematically designed with variations of the chain length of the alcohol and the acid as well as with varying content of the aromatic constituent terephthalic acid (Ta). HPLC/MS identification and quantification of the hydrolysis products terephthalic acid (Ta), benzoic acid (Ba), adipic acid (Ada), mono(4-hydroxybutyl) terephthalate (BTa), mono-(2-hydroxyethyl) terephthalate (ETa), mono-(6-hydroxyhexyl) terephthalate (HTa) and bis(4-hydroxybutyl) terephthalate (BTaB) indicated that these enzymes indeed hydrolyze the tested esters. Shorter terminal chain length acids but longer chain length alcohols in oligomeric model substrates were generally hydrolyzed more efficiently. Thc_Cut1 hydrolyzed aromatic ester bonds more efficiently than HiC resulting in up to 3-fold higher concentrations of the monomeric hydrolysis product Ta. Nevertheless, HiC exhibited a higher overall hydrolytic activity on the tested polyesters, resulting in 2-fold higher concentration of released molecules. Thermogravimetry and differential scanning calorimetry (TG-DSC) of the polymeric model substrates revealed a general trend that a lower difference between melting temperature (Tm) and the temperature at which the enzymatic degradation takes place resulted in higher susceptibility to enzymatic hydrolysis.
ESTHER : Perz_2016_N.Biotechnol_33_295
PubMedSearch : Perz_2016_N.Biotechnol_33_295
PubMedID: 26594021
Gene_locus related to this paper: humin-cut , thefu-q6a0i4

Title : Hydrolysis of synthetic polyesters by Clostridium botulinum esterases - Perz_2016_Biotechnol.Bioeng_113_1024
Author(s) : Perz V , Baumschlager A , Bleymaier K , Zitzenbacher S , Hromic A , Steinkellner G , Pairitsch A , Lyskowski A , Gruber K , Sinkel C , Kuper U , Ribitsch D , Guebitz GM
Ref : Biotechnol Bioeng , 113 :1024 , 2016
Abstract : Two novel esterases from the anaerobe Clostridium botulinum ATCC 3502 (Cbotu_EstA and Cbotu_EstB) were expressed in Escherichia coli BL21-Gold(DE3) and were found to hydrolyze the polyester poly(butylene adipate-co-butylene terephthalate) (PBAT). The active site residues (triad Ser, Asp, His) are present in both enzymes at the same location only with some amino acid variations near the active site at the surrounding of aspartate. Yet, Cbotu_EstA showed higher kcat values on para-nitrophenyl butyrate and para-nitrophenyl acetate and was considerably more active (sixfold) on PBAT. The entrance to the active site of the modeled Cbotu_EstB appears more narrowed compared to the crystal structure of Cbotu_EstA and the N-terminus is shorter which could explain its lower activity on PBAT. The Cbotu_EstA crystal structure consists of two regions that may act as movable cap domains and a zinc metal binding site. Biotechnol. Bioeng. 2016;113: 1024-1034. (c) 2015 Wiley Periodicals, Inc.
ESTHER : Perz_2016_Biotechnol.Bioeng_113_1024
PubMedSearch : Perz_2016_Biotechnol.Bioeng_113_1024
PubMedID: 26524601
Gene_locus related to this paper: clobh-A5I3I2 , clobh-A51055

Title : Biomimetic Approach to Enhance Enzymatic Hydrolysis of the Synthetic Polyester Poly(1,4-butylene adipate): Fusing Binding Modules to Esterases - Perz_2015_Biomacromolecules_16_3889
Author(s) : Perz V , Zumstein MT , Sander M , Zitzenbacher S , Ribitsch D , Guebitz GM
Ref : Biomacromolecules , 16 :3889 , 2015
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.
ESTHER : Perz_2015_Biomacromolecules_16_3889
PubMedSearch : Perz_2015_Biomacromolecules_16_3889
PubMedID: 26566664
Gene_locus related to this paper: thefu-q6a0i4