Author Report for: Zimmermann WNo contact information in database for Zimmermann W
Blazquez-Sanchez P, Vargas JA, Furtado AA, Grinen A, Leonardo DA, Sculaccio SA, Pereira HD, Sonnendecker C, Zimmermann W and Ramirez-Sarmiento CA <2 more author(s)> Blazquez-Sanchez P, Vargas JA, Furtado AA, Grinen A, Leonardo DA, Sculaccio SA, Pereira HD, Sonnendecker C, Zimmermann W, Diez B, Garratt RC, Ramirez-Sarmiento CA (- 2) Ref: Protein Science, :e4757, 2023 : PubMed ESTHER: Blazquez-Sanchez_2023_Protein.Sci__e4757 PubMedSearch: Blazquez-Sanchez 2023 Protein.Sci e4757 PubMedID: 37574805 Gene_locus related to this paper: morsp-lip1 Abstract Several hydrolases have been described to degrade polyethylene terephthalate (PET) at moderate temperatures ranging from 25 degreesC to 40 degreesC. These mesophilic PET hydrolases (PETases) are less efficient in degrading this plastic polymer than their thermophilic homologs, and have therefore been the subject of many protein engineering campaigns. However, enhancing their enzymatic activity through rational design or directed evolution poses a formidable challenge due to the need for exploring a large number of mutations. Additionally, evaluating the improvements in both activity and stability requires screening numerous variants, either individually or using high-throughput screening methods. Here, we utilize instead the design of chimeras as a protein engineering strategy to increase the activity and stability of Mors1, an Antarctic PETase active at 25 degreesC. First, we obtained the crystal structure of Mors1 at 1.6 A resolution, which we used as a scaffold for structure- and sequence-based chimeric design. Then, we designed a Mors1 chimera via loop exchange of a highly divergent active site loop from the thermophilic leaf-branch compost cutinase (LCC) into the equivalent region in Mors1. After restitution of an active site disulfide bond into this chimera, the enzyme exhibited a shift in optimal temperature for activity to 45 degreesC and an increase in 5-fold in PET hydrolysis when compared to wild-type Mors1 at 25 degreesC. Our results serve as a proof of concept of the utility of chimeric design to further improve the activity and stability of PETases active at moderate temperatures. This article is protected by copyright. All rights reserved. Title: On the Binding Mode and Molecular Mechanism of Enzymatic Polyethylene Terephthalate Degradations Falkenstein P, Zhao Z, Di Pede-Mattatelli A, Kunze G, Sommer M, Sonnendecker C, Zimmermann W, Colizzi F, Matysik J, Song C Ref: ACS Catal, 13:6919, 2023 : PubMed ESTHER: Falkenstein_2023_ACS.Catal_13_6919 PubMedSearch: Falkenstein 2023 ACS.Catal 13 6919 Gene_locus related to this paper: thefu-q6a0i4 Abstract Enzymatic degradation of polyethylene terephthlate (PET) by polyester hydrolases is currently subject to intensive research, as it is considered as a potential eco-friendly recycling method for plastic waste. However, the substrate-binding mode and the molecular mechanism of enzymatic PET hydrolysis are still under intense investigation, and controversial hypotheses have been presented. To help unravel the inherent mechanism of biocatalytic PET degradation at the atomic level, we performed solid-state NMR measurements of a cutinase from Thermobifida fusca (TfCut2) embedded in trehalose glasses together with chemically synthesized, amorphous 13C(O)-labeled oligomeric PET. The resulting ternary enzyme-PET-trehalose glassy system enabled advanced solid-state NMR methods for real-time tracking of the enzymatic PET degradation and the investigation of PET chain dynamics. Combined with enhanced-sampling molecular dynamics simulations, specific enzymesubstrate interactions during the degradation process could also be monitored. Our results demonstrate that the PET chain is first cleaved by TfCut2 in blocks of at least one repeat unit and further to terephthalic acid and ethylene glycol. Moreover, the second step (formation of final hydrolysis products) appears to be rate-limiting in such reactions. The observed dynamic changes and interfacial protein contacts of 13C-labeled PET carbonyl groups suggest that only one PET repeat unit is bound to the enzyme during the degradation process while the rest of the PET chain is only loosely confined to the active site. These results, not accessible by using conventional solution enzyme samples and small nonhydrolyzable substrates, provide a better understanding of the biocatalytic PET degradation mechanism of polyester hydrolases. Title: Structure and function of the metagenomic plastic-degrading polyester hydrolase PHL7 bound to its product Richter PK, Blazquez-Sanchez P, Zhao Z, Engelberger F, Wiebeler C, Kunze G, Frank R, Krinke D, Frezzotti E and Sonnendecker C <5 more author(s)> Richter PK, Blazquez-Sanchez P, Zhao Z, Engelberger F, Wiebeler C, Kunze G, Frank R, Krinke D, Frezzotti E, Lihanova Y, Falkenstein P, Matysik J, Zimmermann W, Strater N, Sonnendecker C (- 5) Ref: Nat Commun, 14:1905, 2023 : PubMed ESTHER: Richter_2023_Nat.Commun_14_1905 PubMedSearch: Richter 2023 Nat.Commun 14 1905 PubMedID: 37019924 Gene_locus related to this paper: 9firm-PHL7 Inhibitor(s) related to this paper: Terephthalic-acid Abstract The recently discovered metagenomic-derived polyester hydrolase PHL7 is able to efficiently degrade amorphous polyethylene terephthalate (PET) in post-consumer plastic waste. We present the cocrystal structure of this hydrolase with its hydrolysis product terephthalic acid and elucidate the influence of 17 single mutations on the PET-hydrolytic activity and thermal stability of PHL7. The substrate-binding mode of terephthalic acid is similar to that of the thermophilic polyester hydrolase LCC and deviates from the mesophilic IsPETase. The subsite I modifications L93F and Q95Y, derived from LCC, increased the thermal stability, while exchange of H185S, derived from IsPETase, reduced the stability of PHL7. The subsite II residue H130 is suggested to represent an adaptation for high thermal stability, whereas L210 emerged as the main contributor to the observed high PET-hydrolytic activity. Variant L210T showed significantly higher activity, achieving a degradation rate of 20 microm h(-1) with amorphous PET films. Title: Real-Time Noninvasive Analysis of Biocatalytic PET Degradation Frank R, Krinke D, Sonnendecker C, Zimmermann W, Jahnke HG Ref: ACS Catal, 12:25, 2022 : PubMed ESTHER: Frank_2022_ACS.Catal_12_25 PubMedSearch: Frank 2022 ACS.Catal 12 25 Gene_locus related to this paper: 9firm-PHL7, 9bact-g9by57 Abstract The Earth has entered the Anthropocene, which is branded by ubiquitous and devastating environmental pollution from plastics such as polyethylene terephthalate (PET). Ecofriendly and at the same time economical solutions for plastic recycling and reuse are being sought more urgently now than ever. With the possibility to recover its building blocks, the hydrolysis of PET waste by its selective biodegradation with polyester hydrolases is an appealing solution. We demonstrate how changing the dielectric properties of PET films can be used to evaluate the performance of polyester hydrolases. For this purpose, a PET film separates two reaction chambers in an impedimetric setup to quantify the film thickness- and surface area-dependent change in capacitance caused by the enzyme. The derived degradation rates determined for the polyester hydrolases PHL7 and LCC were similar to those obtained by gravimetric and vertical scanning interferometry measurements. Compared to optical methods, this technique is also insensitive to changes in the solution composition. AFM and FEM simulations further supported that impedance spectroscopy is a powerful tool for the detailed analysis of the enzymatic hydrolysis process of PET films. The developed monitoring system enabled both high-temporal resolution and parallel processing suitable for the analysis of the enzymatic degradability of polyester films and the properties of the biocatalysts. Title: Low Carbon Footprint Recycling of Post-Consumer PET Plastic with a Metagenomic Polyester Hydrolase Sonnendecker C, Oeser J, Richter PK, Hille P, Zhao Z, Fischer C, Lippold H, Blazquez-Sanchez P, Engelberger F and Zimmermann W <9 more author(s)> Sonnendecker C, Oeser J, Richter PK, Hille P, Zhao Z, Fischer C, Lippold H, Blazquez-Sanchez P, Engelberger F, Ramirez-Sarmiento CA, Oeser T, Lihanova Y, Frank R, Jahnke HG, Billig S, Abel B, Strater N, Matysik J, Zimmermann W (- 9) Ref: ChemSusChem, 15:e202101062, 2022 : PubMed ESTHER: Sonnendecker_2022_ChemSusChem_15_e202101062 PubMedSearch: Sonnendecker 2022 ChemSusChem 15 e202101062 PubMedID: 34129279 Gene_locus related to this paper: 9firm-PHL7 Abstract Our planet is flooded with plastics and the need for sustainable recycling strategies of polymers has become increasingly urgent. Enzyme-based hydrolysis of post-consumer plastic is an emerging strategy for closed-loop recycling of polyethylene terephthalate (PET). The polyester hydrolase PHL7 isolated from a compost metagenome completely hydrolyzed amorphous PET films, releasing 91 mg of terephthalic acid per hour and mg of enzyme. Degradation rates of the PET film of 6.8 microm h -1 were monitored by vertical scanning interferometry. Structural analysis indicated the importance of leucine at position 210 for the extraordinarily high PET-hydrolyzing activity of PHL7. Within 24 h, 0.6 mg enzyme g PET -1 completely degraded post-consumer thermoform PET packaging in an aqueous buffer at 70 degreesC without any energy-intensive pretreatments. Terephthalic acid recovered from the enzymatic hydrolysate was used to synthesize virgin PET, demonstrating the potential of polyester hydrolases as catalysts in sustainable PET recycling processes with a low carbon footprint. Title: Antarctic polyester hydrolases degrade aliphatic and aromatic polyesters at moderate temperatures Blazquez-Sanchez P, Engelberger F, Cifuentes-Anticevic J, Sonnendecker C, Grinen A, Reyes J, Diez B, Guixe V, Richter PK and Ramirez-Sarmiento CA <1 more author(s)> Blazquez-Sanchez P, Engelberger F, Cifuentes-Anticevic J, Sonnendecker C, Grinen A, Reyes J, Diez B, Guixe V, Richter PK, Zimmermann W, Ramirez-Sarmiento CA (- 1) Ref: Applied Environmental Microbiology, :AEM0184221, 2021 : PubMed ESTHER: Blazquez-Sanchez_2021_Appl.Environ.Microbiol__AEM0184221 PubMedSearch: Blazquez-Sanchez 2021 Appl.Environ.Microbiol AEM0184221 PubMedID: 34705547 Gene_locus related to this paper: olean-r4ykl9, morsp-lip1 Abstract Polyethylene terephthalate (PET) is one of the most widely used synthetic plastics in the packaging industry, and consequently has become one of the main components of plastic waste found in the environment. However, several microorganisms have been described to encode enzymes that catalyze the depolymerization of PET. While most known PET hydrolases are thermophilic and require reaction temperatures between 60 degreesC to 70 degreesC for an efficient hydrolysis of PET, a partial hydrolysis of amorphous PET at lower temperatures by the polyester hydrolase IsPETase from the mesophilic bacterium Ideonella sakaiensis has also been reported. We show that polyester hydrolases from the Antarctic bacteria Moraxella sp. strain TA144 (Mors1) and Oleispira antarctica RB-8 (OaCut) were able to hydrolyze the aliphatic polyester polycaprolactone as well as the aromatic polyester PET at a reaction temperature of 25 degreesC. Mors1 caused a weight loss of amorphous PET films and thus constitutes a PET-degrading psychrophilic enzyme. Comparative modelling of Mors1 showed that the amino acid composition of its active site resembled both thermophilic and mesophilic PET hydrolases. Lastly, bioinformatic analysis of Antarctic metagenomic samples demonstrated that members of the Moraxellaceae family carry candidate genes coding for further potential psychrophilic PET hydrolases. IMPORTANCE A myriad of consumer products contains polyethylene terephthalate (PET), a plastic that has accumulated as waste in the environment due to its long-term stability and poor waste management. One promising solution is the enzymatic biodegradation of PET, with most known enzymes only catalyzing this process at high temperatures. Here, we bioinformatically identified and biochemically characterized an enzyme from an Antarctic organism that degrades PET at 25 degreesC with similar efficiency than the few PET-degrading enzymes active at moderate temperatures. Reasoning that Antarctica harbors other PET-degrading enzymes, we analyzed available data from Antarctic metagenomic samples and successfully identified other potential enzymes. Our findings contribute to increasing the repertoire of known PET-degrading enzymes that are currently being considered as biocatalysts for the biological recycling of plastic waste. Title: Conformational fitting of a flexible oligomeric substrate does not explain the enzymatic PET degradation Wei R, Song C, Grasing D, Schneider T, Bielytskyi P, Bottcher D, Matysik J, Bornscheuer UT, Zimmermann W Ref: Nat Commun, 10:5581, 2019 : PubMed Title: Fast turbidimetric assay for analyzing the enzymatic hydrolysis of polyethylene terephthalate model substrates Belisario-Ferrari MR, Wei R, Schneider T, Honak A, Zimmermann W Ref: Biotechnol J, :e1800272, 2018 : PubMed ESTHER: Belisario-Ferrari_2018_Biotechnol.J__e1800272 PubMedSearch: Belisario-Ferrari 2018 Biotechnol.J e1800272 PubMedID: 30430764 Gene_locus related to this paper: thefu-1831 Abstract Synthetic plastics such as polyethylene terephthalate (PET) can be cooperatively degraded by microbial polyester hydrolases and carboxylesterases, with the latter hydrolyzing the low-molecular-weight degradation intermediates. For the identification of PET-degrading enzymes, efficient and rapid screening assays are required. Here we report a novel turbidimetric method in a microplate format for the fast screening of enzyme activities against the PET model substrates with two ester bonds bis-(2-Hydroxyethyl) terephthalate (BHET) and ethylene glycol bis-(p-methylbenzoate) (2PET). The carboxylesterase TfCa from Thermobifida fusca KW3 was used for validating the method. High correlation and regression coefficients between the experimental and fitted data confirmed the accuracy and reproducibility of the method and its feasibility for analyzing the kinetics of the enzymatic hydrolysis of the PET model substrates. A comparison of the hydrolysis of BHET and 2PET by TfCa using a kinetic model for heterogeneous catalysis indicated that the enzyme preferentially hydrolyzed the less bulky molecule BHET. The high-throughput assay will facilitate the detection of novel enzymes for the biocatalytic modification or degradation of PET. Title: New insights into the function and global distribution of polyethylene terephthalate (PET) degrading bacteria and enzymes in marine and terrestrial metagenomes Danso D, Schmeisser C, Chow J, Zimmermann W, Wei R, Leggewie C, Li X, Hazen T, Streit WR Ref: Applied Environmental Microbiology, 84:e2773, 2018 : PubMed ESTHER: Danso_2018_Appl.Environ.Microbiol_84_e2773 PubMedSearch: Danso 2018 Appl.Environ.Microbiol 84 e2773 PubMedID: 29427431 Gene_locus related to this paper: 9burk-PET10, 9burk-PET11, 9gamm-a0a0d4l7e6, 9alte-n6vy44, 9zzzz-a0a0f9x315, deiml-e8u721, olean-r4ykl9, vibga-a0a1z2siq1, 9burk-a0a0g3bi90, 9bact-c3ryl0, 9actn-h6wx58, idesa-peth, 9bact-g9by57, acide-PBSA, morsp-lip1 Inhibitor(s) related to this paper: Terephthalic-acid, MHET Abstract Polyethylene terephthalate (PET) is one of the most important synthetic polymers used nowadays. Unfortunately, the polymers accumulate in nature and until now, no highly active enzymes are known that can degrade it at high velocity. Enzymes involved in PET degradation are mainly alpha/beta-hydrolases like cutinases and related enzymes (E.C. 3.1.-). Currently, only a small number of such enzymes are well characterized. Within this work, a search algorithm was developed that identified 504 possible PET hydrolase candidate genes from various databases. A further global search that comprised more than 16 GB of sequence information within 108 marine and 25 terrestrial metagenomes obtained from the IMG data base detected 349 putative PET hydrolases. Heterologous expression of four such candidate enzymes verified the function of these enzymes and confirmed the usefulness of the developed search algorithm. Thereby, two novel and thermostable enzymes with high potential for downstream application were in part characterized. Clustering of 504 novel enzyme candidates based on amino acid similarities indicated that PET hydrolases mainly occur in the phylum of Actinobacteria, Proteobacteria and Bacteroidetes Within the Proteobacteria, the Beta-, Delta- and Gammaproteobacteria were the main hosts. Remarkably enough, in the marine environment, bacteria affiliated with the phylum of the Bacteroidetes appear to be the main host of PET hydrolase genes rather than Actinobacteria or Proteobacteria as observed for the terrestrial metagenomes. Our data further imply that PET hydrolases are truly rare enzymes. The highest occurrence of 1.5 hits/Mb was observed in a sample site containing crude oil.IMPORTANCE Polyethylene terephthalate (PET) accumulates in our environment without significant microbial conversion. Although few PET hydrolases are already known it is still unknown how frequent they appear and which main bacterial phyla they are affiliated with. In this study, deep sequence mining of protein databases and metagenomes demonstrated that PET hydrolases indeed are occurring at very low frequencies in the environment. Further it was possible to link them to phyla which were previously unknown to harbor such enzymes. This work contributes novel knowledge to the phylogenetic relationship, the recent evolution and the global distribution of PET hydrolases. Finally, we describe biochemical traits of four novel PET hydrolases. Title: Degradation of Polyester Polyurethane by Bacterial Polyester Hydrolases Schmidt J, Wei R, Oeser T, Dedavid e Silva LA, Breite D, Schulze A, Zimmermann W Ref: Polymers (Basel), 9:65, 2017 : PubMed ESTHER: Schmidt_2017_Polymers.(Basel)_9_65 PubMedSearch: Schmidt 2017 Polymers.(Basel) 9 65 PubMedID: 30970745 Gene_locus related to this paper: thecd-d1a9g5, thecd-d1a2h1, 9bact-g9by57, thefu-q6a0i4, thefu-q6a0i3 Abstract Polyurethanes (PU) are widely used synthetic polymers. The growing amount of PU used industrially has resulted in a worldwide increase of plastic wastes. The related environmental pollution as well as the limited availability of the raw materials based on petrochemicals requires novel solutions for their efficient degradation and recycling. The degradation of the polyester PU Impranil DLN by the polyester hydrolases LC cutinase (LCC), TfCut2, Tcur1278 and Tcur0390 was analyzed using a turbidimetric assay. The highest hydrolysis rates were obtained with TfCut2 and Tcur0390. TfCut2 also showed a significantly higher substrate affinity for Impranil DLN than the other three enzymes, indicated by a higher adsorption constant K. Significant weight losses of the solid thermoplastic polyester PU (TPU) Elastollan B85A-10 and C85A-10 were detected as a result of the enzymatic degradation by all four polyester hydrolases. Within a reaction time of 200 h at 70 degreesC, LCC caused weight losses of up to 4.9% and 4.1% of Elastollan B85A-10 and C85A-10, respectively. Gel permeation chromatography confirmed a preferential degradation of the larger polymer chains. Scanning electron microscopy revealed cracks at the surface of the TPU cubes as a result of enzymatic surface erosion. Analysis by Fourier transform infrared spectroscopy indicated that the observed weight losses were a result of the cleavage of ester bonds of the polyester TPU. Title: Microbial enzymes for the recycling of recalcitrant petroleum-based plastics: how far are we? Wei R, Zimmermann W Ref: Microb Biotechnol, 10:1308, 2017 : PubMed ESTHER: Wei_2017_Microb.Biotechnol_10_1308 PubMedSearch: Wei 2017 Microb.Biotechnol 10 1308 PubMedID: 28371373 Abstract Petroleum-based plastics have replaced many natural materials in their former applications. With their excellent properties, they have found widespread uses in almost every area of human life. However, the high recalcitrance of many synthetic plastics results in their long persistence in the environment, and the growing amount of plastic waste ending up in landfills and in the oceans has become a global concern. In recent years, a number of microbial enzymes capable of modifying or degrading recalcitrant synthetic polymers have been identified. They are emerging as candidates for the development of biocatalytic plastic recycling processes, by which valuable raw materials can be recovered in an environmentally sustainable way. This review is focused on microbial biocatalysts involved in the degradation of the synthetic plastics polyethylene, polystyrene, polyurethane and polyethylene terephthalate (PET). Recent progress in the application of polyester hydrolases for the recovery of PET building blocks and challenges for the application of these enzymes in alternative plastic waste recycling processes will be discussed. Title: Biocatalysis as a green route for recycling the recalcitrant plastic polyethylene terephthalate Wei R, Zimmermann W Ref: Microb Biotechnol, 10:1302, 2017 : PubMed ESTHER: Wei_2017_Microb.Biotechnol_10_1302 PubMedSearch: Wei 2017 Microb.Biotechnol 10 1302 PubMedID: 28401691 Abstract Biocatalysis can enable a closed-loop recycling of post-consumer PET waste. Title: A dual enzyme system composed of a polyester hydrolase and a carboxylesterase enhances the biocatalytic degradation of polyethylene terephthalate films Barth M, Honak A, Oeser T, Wei R, Belisario-Ferrari MR, Then J, Schmidt J, Zimmermann W Ref: Biotechnol J, 11:1082, 2016 : PubMed ESTHER: Barth_2016_Biotechnol.J_11_1082 PubMedSearch: Barth 2016 Biotechnol.J 11 1082 PubMedID: 27214855 Gene_locus related to this paper: 9bact-g9by57, thefu-1831 Abstract TfCut2 from Thermobifida fusca KW3 and the metagenome-derived LC-cutinase are bacterial polyester hydrolases capable of efficiently degrading polyethylene terephthalate (PET) films. Since the enzymatic PET hydrolysis is inhibited by the degradation intermediate mono-(2-hydroxyethyl) terephthalate (MHET), a dual enzyme system consisting of a polyester hydrolase and the immobilized carboxylesterase TfCa from Thermobifida fusca KW3 was employed for the hydrolysis of PET films at 60 degrees C. HPLC analysis of the reaction products obtained after 24 h of hydrolysis showed an increased amount of soluble products with a lower proportion of MHET in the presence of the immobilized TfCa. The results indicated a continuous hydrolysis of the inhibitory MHET by the immobilized TfCa and demonstrated its advantage as a second biocatalyst in combination with a polyester hydrolase for an efficient degradation oft PET films. The dual enzyme system with LC-cutinase produced a 2.4-fold higher amount of degradation products compared to TfCut2 after a reaction time of 24 h confirming the superior activity of his polyester hydrolase against PET films. Title: Effect of Tris, MOPS, and phosphate buffers on the hydrolysis of polyethylene terephthalate films by polyester hydrolases Schmidt J, Wei R, Oeser T, Belisario-Ferrari MR, Barth M, Then J, Zimmermann W Ref: FEBS Open Bio, 6:919, 2016 : PubMed ESTHER: Schmidt_2016_FEBS.Open.Bio_6_919 PubMedSearch: Schmidt 2016 FEBS.Open.Bio 6 919 PubMedID: 27642555 Abstract The enzymatic degradation of polyethylene terephthalate (PET) occurs at mild reaction conditions and may find applications in environmentally friendly plastic waste recycling processes. The hydrolytic activity of the homologous polyester hydrolases LC cutinase (LCC) from a compost metagenome and TfCut2 from Thermobifida fusca KW3 against PET films was strongly influenced by the reaction medium buffers tris(hydroxymethyl)aminomethane (Tris), 3-(N-morpholino)propanesulfonic acid (MOPS), and sodium phosphate. LCC showed the highest initial hydrolysis rate of PET films in 0.2 m Tris, while the rate of TfCut2 was 2.1-fold lower at this buffer concentration. At a Tris concentration of 1 m, the hydrolysis rate of LCC decreased by more than 90% and of TfCut2 by about 80%. In 0.2 m MOPS or sodium phosphate buffer, no significant differences in the maximum initial hydrolysis rates of PET films by both enzymes were detected. When the concentration of MOPS was increased to 1 m, the hydrolysis rate of LCC decreased by about 90%. The activity of TfCut2 remained low compared to the increasing hydrolysis rates observed at higher concentrations of sodium phosphate buffer. In contrast, the activity of LCC did not change at different concentrations of this buffer. An inhibition study suggested a competitive inhibition of TfCut2 and LCC by Tris and MOPS. Molecular docking showed that Tris and MOPS interfered with the binding of the polymeric substrate in a groove located at the protein surface. A comparison of the K i values and the average binding energies indicated MOPS as the stronger inhibitor of the both enzymes. Title: A disulfide bridge in the calcium binding site of a polyester hydrolase increases its thermal stability and activity against polyethylene terephthalate Then J, Wei R, Oeser T, Gerdts A, Schmidt J, Barth M, Zimmermann W Ref: FEBS Open Bio, 6:425, 2016 : PubMed ESTHER: Then_2016_FEBS.Open.Bio_6_425 PubMedSearch: Then 2016 FEBS.Open.Bio 6 425 PubMedID: 27419048 Gene_locus related to this paper: thefu-q6a0i4 Abstract Elevated reaction temperatures are crucial for the efficient enzymatic degradation of polyethylene terephthalate (PET). A disulfide bridge was introduced to the polyester hydrolase TfCut2 to substitute its calcium binding site. The melting point of the resulting variant increased to 94.7 degreesC (wild-type TfCut2: 69.8 degreesC) and its half-inactivation temperature to 84.6 degreesC (TfCut2: 67.3 degreesC). The variant D204C-E253C-D174R obtained by introducing further mutations at vicinal residues showed a temperature optimum between 75 and 80 degreesC compared to 65 and 70 degreesC of the wild-type enzyme. The variant caused a weight loss of PET films of 25.0 +/- 0.8% (TfCut2: 0.3 +/- 0.1%) at 70 degreesC after a reaction time of 48 h. The results demonstrate that a highly efficient and calcium-independent thermostable polyester hydrolase can be obtained by replacing its calcium binding site with a disulfide bridge. Title: Engineered bacterial polyester hydrolases efficiently degrade polyethylene terephthalate due to relieved product inhibition Wei R, Oeser T, Schmidt J, Meier R, Barth M, Then J, Zimmermann W Ref: Biotechnol Bioeng, 113:1658, 2016 : PubMed ESTHER: Wei_2016_Biotechnol.Bioeng_113_1658 PubMedSearch: Wei 2016 Biotechnol.Bioeng 113 1658 PubMedID: 26804057 Gene_locus related to this paper: 9bact-g9by57, thefu-q6a0i4 Abstract Recent studies on the enzymatic degradation of synthetic polyesters have shown the potential of polyester hydrolases from thermophilic actinomycetes for modifying or degrading polyethylene terephthalate (PET). TfCut2 from Thermobifida fusca KW3 and LC-cutinase (LCC) isolated from a compost metagenome are remarkably active polyester hydrolases with high sequence and structural similarity. Both enzymes exhibit an exposed active site in a substrate binding groove located at the protein surface. By exchanging selected amino acid residues of TfCut2 involved in substrate binding with those present in LCC, enzyme variants with increased PET hydrolytic activity at 65 degrees C were obtained. The highest activity in hydrolyzing PET films and fibers were detected with the single variant G62A and the double variant G62A/I213S. Both variants caused a weight loss of PET films of more than 42% after 50 h of hydrolysis, corresponding to a 2.7-fold increase compared to the wild type enzyme. Kinetic analysis based on the released PET hydrolysis products confirmed the superior hydrolytic activity of G62A with a fourfold higher hydrolysis rate constant and a 1.5-fold lower substrate binding constant than those of the wild type enzyme. Mono-(2-hydroxyethyl) terephthalate is a strong inhibitor of TfCut2. A determination of the Rosetta binding energy suggested a reduced interaction of G62A with 2PET, a dimer of the PET monomer ethylene terephthalate. Indeed, G62A revealed a 5.5-fold lower binding constant to the inhibitor than the wild type enzyme indicating that its increased PET hydrolysis activity is the result of a relieved product inhibition by mono-(2-hydroxyethyl) terephthalate. Biotechnol. Bioeng. 2016;113: 1658-1665. (c) 2016 Wiley Periodicals, Inc. Title: Ca2+ and Mg2+ binding site engineering increases the degradation of polyethylene terephthalate films by polyester hydrolases from Thermobifida fusca Then J, Wei R, Oeser T, Barth M, Belisario-Ferrari MR, Schmidt J, Zimmermann W Ref: Biotechnol J, 10:592, 2015 : PubMed ESTHER: Then_2015_Biotechnol.J_10_592 PubMedSearch: Then 2015 Biotechnol.J 10 592 PubMedID: 25545638 Abstract Several bacterial polyester hydrolases are able to hydrolyze the synthetic polyester polyethylene terephthalate (PET). For an efficient enzymatic degradation of PET, reaction temperatures close to the glass transition temperature of the polymer need to be applied. The esterases TfH, BTA2, Tfu_0882, TfCut1, and TfCut2 produced by the thermophilic actinomycete Thermobifida fusca exhibit PET-hydrolyzing activity. However, these enzymes are not sufficiently stable in this temperature range for an efficient degradation of post-consumer PET materials. The addition of Ca2+ or Mg2+ cations to the enzymes resulted in an increase of their melting points between 10.8 and 14.1 degreesC determined by circular dichroism spectroscopy. The thermostability of the polyester hydrolases was sufficient to degrade semi-crystalline PET films at 65 degreesC in the presence of 10 mM Ca2+ and 10 mM Mg2+ resulting in weight losses of up to 12.9% after a reaction time of 48 h. The residues Asp174, Asp204, and Glu253 were identified by molecular dynamics simulations as potential binding residues for the two cations in TfCut2. This was confirmed by their substitution with arginine, resulting in a higher thermal stability of the corresponding enzyme variants. The generated variants of TfCut2 represent stabilized catalysts suitable for PET hydrolysis reactions performed in the absence of Ca2+ or Mg2+. Title: Structural and functional studies on a thermostable polyethylene terephthalate degrading hydrolase from Thermobifida fusca Roth C, Wei R, Oeser T, Then J, Follner C, Zimmermann W, Strater N Ref: Applied Microbiology & Biotechnology, 98:7815, 2014 : PubMed ESTHER: Roth_2014_Appl.Microbiol.Biotechnol_98_7815 PubMedSearch: Roth 2014 Appl.Microbiol.Biotechnol 98 7815 PubMedID: 24728714 Gene_locus related to this paper: thefu-q6a0i4 Abstract Bacterial cutinases are promising catalysts for the modification and degradation of the widely used plastic polyethylene terephthalate (PET). The improvement of the enzyme for industrial purposes is limited due to the lack of structural information for cutinases of bacterial origin. We have crystallized and structurally characterized a cutinase from Thermobifida fusca KW3 (TfCut2) in free as well as in inhibitor-bound form. Together with our analysis of the thermal stability and modelling studies, we suggest possible reasons for the outstanding thermostability in comparison to the less thermostable homolog from Thermobifida alba AHK119 and propose a model for the binding of the enzyme towards its polymeric substrate. The TfCut2 structure is the basis for the rational design of catalytically more efficient enzyme variants for the hydrolysis of PET and other synthetic polyesters. Title: Functional characterization and structural modeling of synthetic polyester-degrading hydrolases from Thermomonospora curvata Wei R, Oeser T, Then J, Kuhn N, Barth M, Schmidt J, Zimmermann W Ref: AMB Express, 4:44, 2014 : PubMed ESTHER: Wei_2014_AMB.Express_4_44 PubMedSearch: Wei 2014 AMB.Express 4 44 PubMedID: 25405080 Gene_locus related to this paper: thecd-d1a9g5, thecd-d1a2h1 Abstract Thermomonospora curvata is a thermophilic actinomycete phylogenetically related to Thermobifida fusca that produces extracellular hydrolases capable of degrading synthetic polyesters. Analysis of the genome of T. curvata DSM43183 revealed two genes coding for putative polyester hydrolases Tcur1278 and Tcur0390 sharing 61% sequence identity with the T. fusca enzymes. Mature proteins of Tcur1278 and Tcur0390 were cloned and expressed in Escherichia coli TOP10. Tcur1278 and Tcur0390 exhibited an optimal reaction temperature against p-nitrophenyl butyrate at 60 degrees C and 55 degrees C, respectively. The optimal pH for both enzymes was determined at pH 8.5. Tcur1278 retained more than 80% and Tcur0390 less than 10% of their initial activity following incubation for 60 min at 55 degrees C. Tcur0390 showed a higher hydrolytic activity against poly(epsilon-caprolactone) and polyethylene terephthalate (PET) nanoparticles compared to Tcur1278 at reaction temperatures up to 50 degrees C. At 55 degrees C and 60 degrees C, hydrolytic activity against PET nanoparticles was only detected with Tcur1278. In silico modeling of the polyester hydrolases and docking with a model substrate composed of two repeating units of PET revealed the typical fold of alpha/beta serine hydrolases with an exposed catalytic triad. Molecular dynamics simulations confirmed the superior thermal stability of Tcur1278 considered as the main reason for its higher hydrolytic activity on PET. Title: Synthetic polyester-hydrolyzing enzymes from thermophilic actinomycetes Wei R, Oeser T, Zimmermann W Ref: Adv Appl Microbiol, 89:267, 2014 : PubMed ESTHER: Wei_2014_Adv.Appl.Microbiol_89_267 PubMedSearch: Wei 2014 Adv.Appl.Microbiol 89 267 PubMedID: 25131405 Abstract Thermophilic actinomycetes produce enzymes capable of hydrolyzing synthetic polyesters such as polyethylene terephthalate (PET). In addition to carboxylesterases, which have hydrolytic activity predominantly against PET oligomers, esterases related to cutinases also hydrolyze synthetic polymers. The production of these enzymes by actinomycetes as well as their recombinant expression in heterologous hosts is described and their catalytic activity against polyester substrates is compared. Assays to analyze the enzymatic hydrolysis of synthetic polyesters are evaluated, and a kinetic model describing the enzymatic heterogeneous hydrolysis process is discussed. Structure-function and structure-stability relationships of actinomycete polyester hydrolases are compared based on molecular dynamics simulations and recently solved protein structures. In addition, recent progress in enhancing their activity and thermal stability by random or site-directed mutagenesis is presented. Title: Turbidimetric analysis of the enzymatic hydrolysis of polyethylene terephthalate nanoparticles Wei R, Oeser O, Barth M, Weigl N, Luebs A, Schulz-Siegmund M, Hacker MC, Zimmermann W Ref: J Mol Catal B Enzym, 103:72, 2014 : PubMed ESTHER: Wei_2014_J.Mol.Catal.B.Enzym_103_72 PubMedSearch: Wei 2014 J.Mol.Catal.B.Enzym 103 72 Abstract The heterogeneous enzymatic hydrolysis of polyethylene terephthalate (PET) by a polyester hydrolase (TfCut2) from Thermobifida fusca KW3 was determined by measuring the change of intensity of transmitted light due to the scattering effect of PET nanoparticles immobilized in an agarose gel. Nanoparticles with a mean diameter between 100 and 160 nm were prepared from PET samples of different crystallinity to provide a large surface area for the adsorption of the enzyme. The turbidity decrease of the PET nanoparticle suspensions was correlated to the surface erosion process resulting from the enzymatic degradation, and enabled a direct estimation of the kinetic parameters of the enzymatic hydrolysis of PET based on a model for heterogeneous biocatalysis. A comparison of the hydrolysis rate constants and the adsorption equilibrium constants of the enzymatic hydrolysis of PET nanoparticles prepared from recycled PET granulate, film and fibres showed that the biodegradability of PET was mainly influenced by the mobility of the polyester chains, which determined the affinity and accessibility of the ester bonds to the enzyme. Differential scanning calorimetric analysis of the partially hydrolyzed PET nanoparticles provided indirect evidence for an endo-type hydrolytic mechanism of TfCut2 in the heterogeneous degradation of aromatic polyesters. Title: Sugar ester synthesis by thermostable lipase from Streptomyces thermocarboxydus ME168 H-Kittikun A, Prasertsan P, Zimmermann W, Seesuriyachan P, Chaiyaso T Ref: Appl Biochem Biotechnol, 166:1969, 2012 : PubMed ESTHER: H-Kittikun_2012_Appl.Biochem.Biotechnol_166_1969 PubMedSearch: H-Kittikun 2012 Appl.Biochem.Biotechnol 166 1969 PubMedID: 22434352 Abstract The extracellular lipase from Streptomyces thermocarboxydus ME168 was purified to 9.5-fold with 20% yield, following concentration by acetone precipitation, ion exchange chromatography (Resource Q) and gel filtration chromatography (Superdex 200), respectively. The purified enzyme had an apparent molecular mass of 21 kDa by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). The N-terminal sequence of the lipase was ASDFDDQILG and was different from most other reported lipase. The enzyme showed maximum activity at 50 degrees C with the half-life of 180 min at 65 degrees C. It showed high stability at a broad pH range of 5.5-9.5 and was thermostable at the temperature range of 25-60 degrees C. The K(m) and V(max) were 0.28 mM and 1,428 U/mg, respectively, using p-nitrophenyl palmitate as substrate. It was active toward p-nitrophenyl ester with medium to long acyl chain (C(8)-C(16)). Lipase activity was inhibited by Zn(2+), dithiothreitol (DTT), EDTA and some organic solvents, e.g., ethanol, acetone, dioxane, acetronitrile, tert-butanol and pyridine. Immobilized crude lipase of S. thermocarboxydus ME168 on celite could be used to synthesize sugar esters from glucose and vinyl acetate, vinyl butyrate or vinyl caproate in tert-butanol:pyridine (55:45 v/v) at 45 degrees C with conversion yields of 93, 67 and 55%, respectively. Title: Enzymatic Surface Hydrolysis of PET: Effect of Structural Diversity on Kinetic Properties of Cutinases from Thermobifida. Herrero Acero E, Ribitsch D, Steinkellner G, Gruber K, Greimel K, Eiteljoerg I, Trotscha E, Wei R, Zimmermann W and Guebitz G <4 more author(s)> Herrero Acero E, Ribitsch D, Steinkellner G, Gruber K, Greimel K, Eiteljoerg I, Trotscha E, Wei R, Zimmermann W, Zinn M, Cavaco-Paulo A, Freddi G, Schwab H, Guebitz G (- 4) Ref: Macromolecules, 44:4632, 2011 : PubMed ESTHER: Herrero Acero_2011_Macromolecules_44_4632 PubMedSearch: Herrero Acero 2011 Macromolecules 44 4632 Gene_locus related to this paper: bacsu-pnbae, thefu-q6a0i4, thefu-q6a0i3 Abstract In this study cutinases from Thermobifida cellulosilytica DSM44535 (Thc_Cut1 and Thc_Cut2) and Thermobifida fusca DSM44342 (Thf42_Cut1) hydrolyzing poly(ethylene terephthalate) (PET) were successfully cloned and expressed in E.coli BL21-Gold(DE3). Their ability to hydrolyze PET was compared with other enzymes hydrolyzing natural polyesters, including the PHA depolymerase (ePhaZmcl) from Pseudomonas fluorescens and two cutinases from T. fusca KW3. The three isolated Thermobifida cutinases are very similar (only a maximum of 18 amino acid differences) but yet had different kinetic parameters on soluble substrates. Their kcat and Km values on pNP-acetate were in the ranges 2.4-211.9 s-1 and 127-200 micoM while on pNP-butyrate they showed kcat and Km values between 5.3 and 195.1 s-1 and between 1483 and 2133 microM. Thc_Cut1 released highest amounts of MHET and terephthalic acid from PET and bis(benzoyloxyethyl) terephthalate (3PET) with the highest concomitant increase in PET hydrophilicity as indicated by water contact angle (WCA) decreases. FTIR-ATR analysis revealed an increase in the crystallinity index A1340/A1410 upon enzyme treatment and an increase of the amount of carboxylic and hydroxylic was measured using derivatization with 2-(bromomethyl)naphthalene. Modeling the covalently bound tetrahedral intermediate consisting of cutinase and 3PET indicated that the active site His-209 is in the proximity of the O of the substrate thus allowing hydrolysis. On the other hand, the models indicated that regions of Thc_Cut1 and Thc_Cut2 which differed in electrostatic and in hydrophobic surface properties were able to reach/interact with PET which may explain their different hydrolysis efficiencies. Title: Enzymes for the biofunctionalization of poly(ethylene terephthalate) Zimmermann W, Billig S Ref: Adv Biochem Eng Biotechnol, 125:97, 2011 : PubMed ESTHER: Zimmermann_2011_Adv.Biochem.Eng.Biotechnol_125_97 PubMedSearch: Zimmermann 2011 Adv.Biochem.Eng.Biotechnol 125 97 PubMedID: 21076908 Abstract The functionalization of synthetic polymers such as poly(ethylene terephthalate) to improve their hydrophilicity can be achieved biocatalytically using hydrolytic enzymes. A number of cutinases, lipases, and esterases active on polyethylene terephthalate have been identified and characterized. Enzymes from Fusarium solani, Thermomyces insolens, T. lanuginosus, Aspergillus oryzae, Pseudomonas mendocina, and Thermobifida fusca have been studied in detail. Thermostable biocatalysts hydrolyzing poly(ethylene terephthalate) are promising candidates for the further optimization of suitable biofunctionalization processes for textile finishing, technical, and biomedical applications. Title: Hydrolysis of cyclic poly(ethylene terephthalate) trimers by a carboxylesterase from Thermobifida fusca KW3 Billig S, Oeser T, Birkemeyer C, Zimmermann W Ref: Applied Microbiology & Biotechnology, 87:1753, 2010 : PubMed ESTHER: Billig_2010_Appl.Microbiol.Biotechnol_87_1753 PubMedSearch: Billig 2010 Appl.Microbiol.Biotechnol 87 1753 PubMedID: 20467738 Gene_locus related to this paper: thefu-1831 Abstract We have identified a carboxylesterase produced in liquid cultures of the thermophilic actinomycete Thermobifida fusca KW3 that were supplemented with poly(ethylene terephthalate) fibers. The enzyme hydrolyzed highly hydrophobic, synthetic cyclic poly(ethylene terephthalate) trimers with an optimal activity at 60 degrees C and a pH of 6. V (max) and K (m) values for the hydrolysis were 9.3 micromol(-1) min(-1) mg(-1) and 0.5 mM, respectively. The esterase showed high specificity towards short and middle chain-length fatty acyl esters of p-nitrophenol. The enzyme retained 37% of its activity after 96 h of incubation at 50 degrees C and a pH of 8. Enzyme inhibition studies and analysis of substitution mutants of the carboxylesterase revealed the typical catalytic mechanism of a serine hydrolase with a catalytic triad composed of serine, glutamic acid, and histidine. Title: Biochemical characterization of the cutinases from Thermobifida fusca Chen S, Su L, Billig S, Zimmermann W, Chen J, Wu J Ref: J Mol Catal B Enzym, 63:121, 2010 : PubMed ESTHER: Chen_2010_J.Mol.Catal.B.Enzym_63_121 PubMedSearch: Chen 2010 J.Mol.Catal.B.Enzym 63 121 Gene_locus related to this paper: thefu-q6a0i4, thefu-q6a0i3 Abstract Thermobifida fusca produces two cutinases which share 93% identity in amino acid sequence. In the present study, we investigated the detailed biochemical properties of T. fusca cutinases for the first time. For a better comparison between bacterial and fungal cutinases, recombinant Fusarium solani pisi cutinase was subjected to the similar analysis. The results showed that both bacterial and fungal cutinases are monomeric proteins in solution. The bacterial cutinases exhibited a broad substrate specificity against plant cutin, synthetic polyesters, insoluble triglycerides, and soluble esters. In addition, the two isoenzymes of T. fusca and the F.solani pisi cutinase are similar in substrate kinetics, the lack of interfacial activation, and metal ion requirements. However, the T.fusca cutinases showed higher stability in the presence of surfactants and organic solvents. Considering the versatile hydrolytic activity, good tolerance to surfactants, superior stability in organic solvents, and thermostability demonstrated by T. fusca cutinases, they may have promising applications in related industries. Title: The genome of Streptococcus mitis B6--what is a commensal? Denapaite D, Bruckner R, Nuhn M, Reichmann P, Henrich B, Maurer P, Schahle Y, Selbmann P, Zimmermann W and Hakenbeck R <1 more author(s)> Denapaite D, Bruckner R, Nuhn M, Reichmann P, Henrich B, Maurer P, Schahle Y, Selbmann P, Zimmermann W, Wambutt R, Hakenbeck R (- 1) Ref: PLoS ONE, 5:e9426, 2010 : PubMed ESTHER: Denapaite_2010_PLoS.ONE_5_E9426 PubMedSearch: Denapaite 2010 PLoS.ONE 5 E9426 PubMedID: 20195536 Gene_locus related to this paper: strm6-d3h7r7, strm6-d3h9p9, strm6-d3h9q0, strm6-d3hac7, strpn-AXE1, strm6-d3haj3 Abstract Streptococcus mitis is the closest relative of the major human pathogen S. pneumoniae. The 2,15 Mb sequence of the Streptococcus mitis B6 chromosome, an unusually high-level beta-lactam resistant and multiple antibiotic resistant strain, has now been determined to encode 2100 genes. The accessory genome is estimated to represent over 40%, including 75 mostly novel transposases and IS, the prophage phiB6 and another seven phage related regions. Tetracycline resistance mediated by Tn5801, and an unusual and large gene cluster containing three aminoglycoside resistance determinants have not been described in other Streptococcus spp. Comparative genomic analyses including hybridization experiments on a S. mitis B6 specific microarray reveal that individual S. mitis strains are almost as distantly related to the B6 strain as S. pneumoniae. Both species share a core of over 900 genes. Most proteins described as pneumococcal virulence factors are present in S. mitis B6, but the three choline binding proteins PcpA, PspA and PspC, and three gene clusters containing the hyaluronidase gene, ply and lytA, and the capsular genes are absent in S. mitis B6 and other S. mitis as well and confirm their importance for the pathogenetic potential of S. pneumoniae. Despite the close relatedness between the two species, the S. mitis B6 genome reveals a striking X-alignment when compared with S. pneumoniae. Title: High level expression of a hydrophobic poly(ethylene terephthalate)-hydrolyzing carboxylesterase from Thermobifida fusca KW3 in Escherichia coli BL21(DE3) Oeser T, Wei R, Baumgarten T, Billig S, Follner C, Zimmermann W Ref: J Biotechnol, 146:100, 2010 : PubMed ESTHER: Oeser_2010_J.Biotechnol_146_100 PubMedSearch: Oeser 2010 J.Biotechnol 146 100 PubMedID: 20156495 Gene_locus related to this paper: thefu-1831 Abstract The gram-positive thermophilic actinomycete Thermobifida fusca KW3 secretes a highly hydrophobic carboxylesterase (TfCa) that is able to hydrolyze poly(ethylene terephthalate). TfCa was produced in the Escherichia coli strain BL21(DE3) as a fusion protein consisting of a pelB leader sequence to ensure periplasmic localization of the protein and a His(6) tag for use in its purification. To enhance the recombinant enzyme yield, the tfca gene from T. fusca KW3 was successfully optimized for codon usage in E. coli. In addition, the gene expression induction conditions were optimized and the temperature for cell cultivation was lowered to reduce inclusion body formation. The optimized codons and expression conditions yielded 4500-fold higher TfCa activity than the wild-type strain. Using a pH-controlled bioreactor for cultivation, a TfCa protein concentration of 41.6mg/L was achieved. Title: Increase of the hydrophilicity of polyethylene terephthalate fibres by hydrolases from Thermomonospora fusca and Fusarium solani f. sp. pisi Alisch-Mark M, Herrmann A, Zimmermann W Ref: Biotechnol Lett, 28:681, 2006 : PubMed ESTHER: Alisch-Mark_2006_Biotechnol.Lett_28_681 PubMedSearch: Alisch-Mark 2006 Biotechnol.Lett 28 681 PubMedID: 16791721 Abstract Treatment of polyethylene terephthalate fibres with hydrolase preparations from Thermomonospora (Thermobifida) fusca and Fusarium solani f. sp. pisi resulted in an increase of the hydrophilicity of the fibres determined by measurement of their dyeing behaviour with reactive dyes and their water absorption ability. Reflectance spectrometry of treated fibres dyed with a reactive dye showed that the colour became more intense corresponding to an increase of hydroxyl groups on the fibre surfaces and indicated a stepwise peeling of the fibres by the enzymes comparable to the effects obtained by alkaline treatments. The synthetic fibres treated with the hydrolase from T. fusca also showed enhanced water absorption ability further confirming the increased surface hydrophilicity caused by the enzyme. Title: The genome of the kinetoplastid parasite, Leishmania major Ivens AC, Peacock CS, Worthey EA, Murphy L, Aggarwal G, Berriman M, Sisk E, Rajandream MA, Adlem E and Myler PJ <91 more author(s)> Ivens AC, Peacock CS, Worthey EA, Murphy L, Aggarwal G, Berriman M, Sisk E, Rajandream MA, Adlem E, Aert R, Anupama A, Apostolou Z, Attipoe P, Bason N, Bauser C, Beck A, Beverley SM, Bianchettin G, Borzym K, Bothe G, Bruschi CV, Collins M, Cadag E, Ciarloni L, Clayton C, Coulson RM, Cronin A, Cruz AK, Davies RM, De Gaudenzi J, Dobson DE, Duesterhoeft A, Fazelina G, Fosker N, Frasch AC, Fraser A, Fuchs M, Gabel C, Goble A, Goffeau A, Harris D, Hertz-Fowler C, Hilbert H, Horn D, Huang Y, Klages S, Knights A, Kube M, Larke N, Litvin L, Lord A, Louie T, Marra M, Masuy D, Matthews K, Michaeli S, Mottram JC, Muller-Auer S, Munden H, Nelson S, Norbertczak H, Oliver K, O'Neil S, Pentony M, Pohl TM, Price C, Purnelle B, Quail MA, Rabbinowitsch E, Reinhardt R, Rieger M, Rinta J, Robben J, Robertson L, Ruiz JC, Rutter S, Saunders D, Schafer M, Schein J, Schwartz DC, Seeger K, Seyler A, Sharp S, Shin H, Sivam D, Squares R, Squares S, Tosato V, Vogt C, Volckaert G, Wambutt R, Warren T, Wedler H, Woodward J, Zhou S, Zimmermann W, Smith DF, Blackwell JM, Stuart KD, Barrell B, Myler PJ (- 91) Ref: Science, 309:436, 2005 : PubMed ESTHER: Ivens_2005_Science_309_436 PubMedSearch: Ivens 2005 Science 309 436 PubMedID: 16020728 Gene_locus related to this paper: leima-e9ady6, leima-L2464.12, leima-L2802.02, leima-OPB, leima-q4fw33, leima-q4fwg8, leima-q4fwj0, leima-q4fya7, leima-q4q0a1, leima-q4q0t5, leima-q4q0v0, leima-q4q1h9, leima-q4q2c9, leima-q4q4j7, leima-q4q4t6, leima-q4q5j1, leima-q4q6e9, leima-q4q7v8, leima-q4q8a8, leima-q4q9g9, leima-q4q080, leima-q4q398, leima-q4q615, leima-q4q819, leima-q4q871, leima-q4q942, leima-q4qae7, leima-q4qb85, leima-q4qdz7, leima-q4qe26, leima-q4qe31, leima-q4qe85, leima-q4qe86, leima-q4qe87, leima-q4qe90, leima-q4qec8, leima-q4qgz4, leima-q4qgz5, leima-q4qhs0, leima-q4qj45 Abstract Leishmania species cause a spectrum of human diseases in tropical and subtropical regions of the world. We have sequenced the 36 chromosomes of the 32.8-megabase haploid genome of Leishmania major (Friedlin strain) and predict 911 RNA genes, 39 pseudogenes, and 8272 protein-coding genes, of which 36% can be ascribed a putative function. These include genes involved in host-pathogen interactions, such as proteolytic enzymes, and extensive machinery for synthesis of complex surface glycoconjugates. The organization of protein-coding genes into long, strand-specific, polycistronic clusters and lack of general transcription factors in the L. major, Trypanosoma brucei, and Trypanosoma cruzi (Tritryp) genomes suggest that the mechanisms regulating RNA polymerase II-directed transcription are distinct from those operating in other eukaryotes, although the trypanosomatids appear capable of chromatin remodeling. Abundant RNA-binding proteins are encoded in the Tritryp genomes, consistent with active posttranscriptional regulation of gene expression. Title: The genome sequence of Schizosaccharomyces pombe Wood V, Gwilliam R, Rajandream MA, Lyne M, Lyne R, Stewart A, Sgouros J, Peat N, Hayles J and Nurse P <124 more author(s)> Wood V, Gwilliam R, Rajandream MA, Lyne M, Lyne R, Stewart A, Sgouros J, Peat N, Hayles J, Baker S, Basham D, Bowman S, Brooks K, Brown D, Brown S, Chillingworth T, Churcher C, Collins M, Connor R, Cronin A, Davis P, Feltwell T, Fraser A, Gentles S, Goble A, Hamlin N, Harris D, Hidalgo J, Hodgson G, Holroyd S, Hornsby T, Howarth S, Huckle EJ, Hunt S, Jagels K, James K, Jones L, Jones M, Leather S, McDonald S, McLean J, Mooney P, Moule S, Mungall K, Murphy L, Niblett D, Odell C, Oliver K, O'Neil S, Pearson D, Quail MA, Rabbinowitsch E, Rutherford K, Rutter S, Saunders D, Seeger K, Sharp S, Skelton J, Simmonds M, Squares R, Squares S, Stevens K, Taylor K, Taylor RG, Tivey A, Walsh S, Warren T, Whitehead S, Woodward J, Volckaert G, Aert R, Robben J, Grymonprez B, Weltjens I, Vanstreels E, Rieger M, Schafer M, Muller-Auer S, Gabel C, Fuchs M, Dusterhoft A, Fritzc C, Holzer E, Moestl D, Hilbert H, Borzym K, Langer I, Beck A, Lehrach H, Reinhardt R, Pohl TM, Eger P, Zimmermann W, Wedler H, Wambutt R, Purnelle B, Goffeau A, Cadieu E, Dreano S, Gloux S, Lelaure V, Mottier S, Galibert F, Aves SJ, Xiang Z, Hunt C, Moore K, Hurst SM, Lucas M, Rochet M, Gaillardin C, Tallada VA, Garzon A, Thode G, Daga RR, Cruzado L, Jimenez J, Sanchez M, del Rey F, Benito J, Dominguez A, Revuelta JL, Moreno S, Armstrong J, Forsburg SL, Cerutti L, Lowe T, McCombie WR, Paulsen I, Potashkin J, Shpakovski GV, Ussery D, Barrell BG, Nurse P (- 124) Ref: Nature, 415:871, 2002 : PubMed ESTHER: Wood_2002_Nature_415_871 PubMedSearch: Wood 2002 Nature 415 871 PubMedID: 11859360 Gene_locus related to this paper: schpo-APTH1, schpo-be46, schpo-BST1, schpo-C2E11.08, schpo-C14C4.15C, schpo-C22H12.03, schpo-C23C4.16C, schpo-C57A10.08C, schpo-dyr, schpo-este1, schpo-KEX1, schpo-PCY1, schpo-pdat, schpo-PLG7, schpo-ppme1, schpo-q9c0y8, schpo-SPAC4A8.06C, schpo-C22A12.06C, schpo-SPAC977.15, schpo-SPAPB1A11.02, schpo-SPBC14C8.15, schpo-SPBC530.12C, schpo-SPBC1711.12, schpo-SPBPB2B2.02, schpo-SPCC5E4.05C, schpo-SPCC417.12, schpo-SPCC1672.09, schpo-yb4e, schpo-yblh, schpo-ydw6, schpo-ye7a, schpo-ye63, schpo-ye88, schpo-yeld, schpo-yk68, schpo-clr3, schpo-ykv6 Abstract We have sequenced and annotated the genome of fission yeast (Schizosaccharomyces pombe), which contains the smallest number of protein-coding genes yet recorded for a eukaryote: 4,824. The centromeres are between 35 and 110 kilobases (kb) and contain related repeats including a highly conserved 1.8-kb element. Regions upstream of genes are longer than in budding yeast (Saccharomyces cerevisiae), possibly reflecting more-extended control regions. Some 43% of the genes contain introns, of which there are 4,730. Fifty genes have significant similarity with human disease genes; half of these are cancer related. We identify highly conserved genes important for eukaryotic cell organization including those required for the cytoskeleton, compartmentation, cell-cycle control, proteolysis, protein phosphorylation and RNA splicing. These genes may have originated with the appearance of eukaryotic life. Few similarly conserved genes that are important for multicellular organization were identified, suggesting that the transition from prokaryotes to eukaryotes required more new genes than did the transition from unicellular to multicellular organization. Title: The DNA sequence of human chromosome 21 Hattori M, Fujiyama A, Taylor TD, Watanabe H, Yada T, Park HS, Toyoda A, Ishii K, Totoki Y and Yaspo ML <54 more author(s)> Hattori M, Fujiyama A, Taylor TD, Watanabe H, Yada T, Park HS, Toyoda A, Ishii K, Totoki Y, Choi DK, Groner Y, Soeda E, Ohki M, Takagi T, Sakaki Y, Taudien S, Blechschmidt K, Polley A, Menzel U, Delabar J, Kumpf K, Lehmann R, Patterson D, Reichwald K, Rump A, Schillhabel M, Schudy A, Zimmermann W, Rosenthal A, Kudoh J, Schibuya K, Kawasaki K, Asakawa S, Shintani A, Sasaki T, Nagamine K, Mitsuyama S, Antonarakis SE, Minoshima S, Shimizu N, Nordsiek G, Hornischer K, Brant P, Scharfe M, Schon O, Desario A, Reichelt J, Kauer G, Blocker H, Ramser J, Beck A, Klages S, Hennig S, Riesselmann L, Dagand E, Haaf T, Wehrmeyer S, Borzym K, Gardiner K, Nizetic D, Francis F, Lehrach H, Reinhardt R, Yaspo ML (- 54) Ref: Nature, 405:311, 2000 : PubMed ESTHER: Hattori_2000_Nature_405_311 PubMedSearch: Hattori 2000 Nature 405 311 PubMedID: 10830953 Gene_locus related to this paper: human-LIPI Abstract Chromosome 21 is the smallest human autosome. An extra copy of chromosome 21 causes Down syndrome, the most frequent genetic cause of significant mental retardation, which affects up to 1 in 700 live births. Several anonymous loci for monogenic disorders and predispositions for common complex disorders have also been mapped to this chromosome, and loss of heterozygosity has been observed in regions associated with solid tumours. Here we report the sequence and gene catalogue of the long arm of chromosome 21. We have sequenced 33,546,361 base pairs (bp) of DNA with very high accuracy, the largest contig being 25,491,867 bp. Only three small clone gaps and seven sequencing gaps remain, comprising about 100 kilobases. Thus, we achieved 99.7% coverage of 21q. We also sequenced 281,116 bp from the short arm. The structural features identified include duplications that are probably involved in chromosomal abnormalities and repeat structures in the telomeric and pericentromeric regions. Analysis of the chromosome revealed 127 known genes, 98 predicted genes and 59 pseudogenes. Title: Sequence and analysis of chromosome 5 of the plant Arabidopsis thaliana Tabata S, Kaneko T, Nakamura Y, Kotani H, Kato T, Asamizu E, Miyajima N, Sasamoto S, Kimura T and Fransz P <119 more author(s)> Tabata S, Kaneko T, Nakamura Y, Kotani H, Kato T, Asamizu E, Miyajima N, Sasamoto S, Kimura T, Hosouchi T, Kawashima K, Kohara M, Matsumoto M, Matsuno A, Muraki A, Nakayama S, Nakazaki N, Naruo K, Okumura S, Shinpo S, Takeuchi C, Wada T, Watanabe A, Yamada M, Yasuda M, Sato S, de la Bastide M, Huang E, Spiegel L, Gnoj L, O'Shaughnessy A, Preston R, Habermann K, Murray J, Johnson D, Rohlfing T, Nelson J, Stoneking T, Pepin K, Spieth J, Sekhon M, Armstrong J, Becker M, Belter E, Cordum H, Cordes M, Courtney L, Courtney W, Dante M, Du H, Edwards J, Fryman J, Haakensen B, Lamar E, Latreille P, Leonard S, Meyer R, Mulvaney E, Ozersky P, Riley A, Strowmatt C, Wagner-McPherson C, Wollam A, Yoakum M, Bell M, Dedhia N, Parnell L, Shah R, Rodriguez M, See LH, Vil D, Baker J, Kirchoff K, Toth K, King L, Bahret A, Miller B, Marra M, Martienssen R, McCombie WR, Wilson RK, Murphy G, Bancroft I, Volckaert G, Wambutt R, Dusterhoft A, Stiekema W, Pohl T, Entian KD, Terryn N, Hartley N, Bent E, Johnson S, Langham SA, McCullagh B, Robben J, Grymonprez B, Zimmermann W, Ramsperger U, Wedler H, Balke K, Wedler E, Peters S, van Staveren M, Dirkse W, Mooijman P, Lankhorst RK, Weitzenegger T, Bothe G, Rose M, Hauf J, Berneiser S, Hempel S, Feldpausch M, Lamberth S, Villarroel R, Gielen J, Ardiles W, Bents O, Lemcke K, Kolesov G, Mayer K, Rudd S, Schoof H, Schueller C, Zaccaria P, Mewes HW, Bevan M, Fransz P (- 119) Ref: Nature, 408:823, 2000 : PubMed ESTHER: Tabata_2000_Nature_408_823 PubMedSearch: Tabata 2000 Nature 408 823 PubMedID: 11130714 Gene_locus related to this paper: arath-At5g11650, arath-At5g16120, arath-at5g18630, arath-AT5G20520, arath-At5g21950, arath-AT5G27320, arath-CXE15, arath-F1N13.220, arath-F14F8.240, arath-q3e9e4, arath-q8lae9, arath-Q8LFB7, arath-q9ffg7, arath-q9fij5, arath-Q9LVU7, arath-q66gm8, arath-SCPL34, arath-B9DFR3, arath-a0a1p8bcz0 Abstract The genome of the model plant Arabidopsis thaliana has been sequenced by an international collaboration, The Arabidopsis Genome Initiative. Here we report the complete sequence of chromosome 5. This chromosome is 26 megabases long; it is the second largest Arabidopsis chromosome and represents 21% of the sequenced regions of the genome. The sequence of chromosomes 2 and 4 have been reported previously and that of chromosomes 1 and 3, together with an analysis of the complete genome sequence, are reported in this issue. Analysis of the sequence of chromosome 5 yields further insights into centromere structure and the sequence determinants of heterochromatin condensation. The 5,874 genes encoded on chromosome 5 reveal several new functions in plants, and the patterns of gene organization provide insights into the mechanisms and extent of genome evolution in plants. Title: Sequence and analysis of chromosome 4 of the plant Arabidopsis thaliana Mayer K, Schuller C, Wambutt R, Murphy G, Volckaert G, Pohl T, Dusterhoft A, Stiekema W, Entian KD and McCombie WR <220 more author(s)> Mayer K, Schuller C, Wambutt R, Murphy G, Volckaert G, Pohl T, Dusterhoft A, Stiekema W, Entian KD, Terryn N, Harris B, Ansorge W, Brandt P, Grivell L, Rieger M, Weichselgartner M, de Simone V, Obermaier B, Mache R, Muller M, Kreis M, Delseny M, Puigdomenech P, Watson M, Schmidtheini T, Reichert B, Portatelle D, Perez-Alonso M, Boutry M, Bancroft I, Vos P, Hoheisel J, Zimmermann W, Wedler H, Ridley P, Langham SA, McCullagh B, Bilham L, Robben J, Van der Schueren J, Grymonprez B, Chuang YJ, Vandenbussche F, Braeken M, Weltjens I, Voet M, Bastiaens I, Aert R, Defoor E, Weitzenegger T, Bothe G, Ramsperger U, Hilbert H, Braun M, Holzer E, Brandt A, Peters S, van Staveren M, Dirske W, Mooijman P, Klein Lankhorst R, Rose M, Hauf J, Kotter P, Berneiser S, Hempel S, Feldpausch M, Lamberth S, Van den Daele H, De Keyser A, Buysshaert C, Gielen J, Villarroel R, De Clercq R, van Montagu M, Rogers J, Cronin A, Quail M, Bray-Allen S, Clark L, Doggett J, Hall S, Kay M, Lennard N, McLay K, Mayes R, Pettett A, Rajandream MA, Lyne M, Benes V, Rechmann S, Borkova D, Blocker H, Scharfe M, Grimm M, Lohnert TH, Dose S, de Haan M, Maarse A, Schafer M, Muller-Auer S, Gabel C, Fuchs M, Fartmann B, Granderath K, Dauner D, Herzl A, Neumann S, Argiriou A, Vitale D, Liguori R, Piravandi E, Massenet O, Quigley F, Clabauld G, Mundlein A, Felber R, Schnabl S, Hiller R, Schmidt W, Lecharny A, Aubourg S, Chefdor F, Cooke R, Berger C, Montfort A, Casacuberta E, Gibbons T, Weber N, Vandenbol M, Bargues M, Terol J, Torres A, Perez-Perez A, Purnelle B, Bent E, Johnson S, Tacon D, Jesse T, Heijnen L, Schwarz S, Scholler P, Heber S, Francs P, Bielke C, Frishman D, Haase D, Lemcke K, Mewes HW, Stocker S, Zaccaria P, Bevan M, Wilson RK, de la Bastide M, Habermann K, Parnell L, Dedhia N, Gnoj L, Schutz K, Huang E, Spiegel L, Sehkon M, Murray J, Sheet P, Cordes M, Abu-Threideh J, Stoneking T, Kalicki J, Graves T, Harmon G, Edwards J, Latreille P, Courtney L, Cloud J, Abbott A, Scott K, Johnson D, Minx P, Bentley D, Fulton B, Miller N, Greco T, Kemp K, Kramer J, Fulton L, Mardis E, Dante M, Pepin K, Hillier L, Nelson J, Spieth J, Ryan E, Andrews S, Geisel C, Layman D, Du H, Ali J, Berghoff A, Jones K, Drone K, Cotton M, Joshu C, Antonoiu B, Zidanic M, Strong C, Sun H, Lamar B, Yordan C, Ma P, Zhong J, Preston R, Vil D, Shekher M, Matero A, Shah R, Swaby IK, O'Shaughnessy A, Rodriguez M, Hoffmann J, Till S, Granat S, Shohdy N, Hasegawa A, Hameed A, Lodhi M, Johnson A, Chen E, Marra M, Martienssen R, McCombie WR (- 220) Ref: Nature, 402:769, 1999 : PubMed ESTHER: Mayer_1999_Nature_402_769 PubMedSearch: Mayer 1999 Nature 402 769 PubMedID: 10617198 Gene_locus related to this paper: arath-AT4G00500, arath-AT4G16690, arath-AT4G17480, arath-AT4G24380, arath-AT4g30610, arath-o65513, arath-o65713, arath-LPAAT, arath-f4jt64 Abstract The higher plant Arabidopsis thaliana (Arabidopsis) is an important model for identifying plant genes and determining their function. To assist biological investigations and to define chromosome structure, a coordinated effort to sequence the Arabidopsis genome was initiated in late 1996. Here we report one of the first milestones of this project, the sequence of chromosome 4. Analysis of 17.38 megabases of unique sequence, representing about 17% of the genome, reveals 3,744 protein coding genes, 81 transfer RNAs and numerous repeat elements. Heterochromatic regions surrounding the putative centromere, which has not yet been completely sequenced, are characterized by an increased frequency of a variety of repeats, new repeats, reduced recombination, lowered gene density and lowered gene expression. Roughly 60% of the predicted protein-coding genes have been functionally characterized on the basis of their homology to known genes. Many genes encode predicted proteins that are homologous to human and Caenorhabditis elegans proteins. | |||||||
|
