Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

    Full text documents are added when available. The database is updated daily and currently holds about 240,000 items, of which 72,000 in open access.

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Improved protein production and codon optimization analyses in Escherichia coli by bicistronic design
Nieuwkoop, Thijs ; Claassens, Nico J. ; Oost, John van der - \ 2018
Microbial Biotechnology (2018). - ISSN 1751-7907 - 7 p.

Different codon optimization algorithms are available that aim at improving protein production by optimizing translation elongation. In these algorithms, it is generally not considered how the altered protein coding sequence will affect the secondary structure of the corresponding RNA transcript, particularly not the effect on the 5′-UTR structure and related ribosome binding site availability. This is a serious drawback, because the influence of codon usage on mRNA secondary structures, especially near the start of a gene, may strongly influence translation initiation. In this study, we aim to reduce the effect of codon usage on translation initiation by applying a bicistronic design (BCD) element. Protein production of several codon-optimized gene variants is tested in parallel for a BCD and a standard monocistronic design (MCD). We demonstrate that these distinct architectures can drastically change the relative performance of different codon optimization algorithms. We conclude that a BCD is indispensable in future studies that aim to reveal the impact of codon optimization and codon usage correlations. Furthermore, irrespective of the algorithm used, using a BCD does improve protein production compared with an MCD. The overall highest expression from BCDs for both GFP and RFP is at least twofold higher than the highest levels found for the MCDs, while for codon variants having very low expression from the MCD, even 10-fold to 100-fold increases in expression were achieved by the BCD. This shows the great potential of the BCD element for recombinant protein production.

Towards biobased ethyl acetate and beyond : Identification and application of the elusive Eat1 enzyme
Kruis, Aleksander Johannes - \ 2018
Wageningen University. Promotor(en): Ruud Weusthuis; John van der Oost, co-promotor(en): Servé Kengen. - Wageningen : Wageningen University - ISBN 9789463435260 - 251
The alcohol acetyltransferase Eat1 is located in yeast mitochondria
Kruis, A.J. ; Mars, A.E. ; Kengen, S.W.M. ; Borst, J.W. ; Oost, J. van der; Weusthuis, R.A. - \ 2018
Applied and Environmental Microbiology 84 (2018)19. - ISSN 0099-2240 - p. 1 - 11.
Eat1 is a recently discovered alcohol acetyltransferase responsible for bulk ethyl acetate production in yeasts such as Wickerhamomyces anomalus and Kluyveromyces lactis. These yeasts have the potential to become efficient biobased ethyl acetate producers. However, some fundamental features of Eat1 are still not understood, which hampers the rational engineering of efficient production strains. The cellular location of Eat1 in yeast is one of these features. To reveal its location, Eat1 was fused with yEGFP to allow intracellular tracking. Despite the current assumption that bulk ethyl acetate production occurs in the yeast cytosol, most of Eat1 localised to the mitochondria of K. lactis CBS 2359 Δku80. We then compared five bulk ethyl acetate-producing yeasts in iron-limited chemostats with glucose as carbon source. All yeasts produced ethyl acetate under these conditions. This strongly suggests that the mechanism and location of bulk ethyl acetate synthesis are similar in these yeast strains. Furthermore, an in silico analysis showed that Eat1 proteins from various yeasts were mostly predicted as mitochondrial. Altogether, it is concluded that Eat1-catalyzed ethyl acetate production occurs in yeast mitochondria. This study has added new insights to bulk ethyl acetate synthesis in yeast, which is relevant for developing efficient production strains.
Angiopoietin-like 4 promotes the intracellular cleavage of lipoprotein lipase by PCSK3/furin in adipocytes
Dijk, Wieneke ; Ruppert, Philip M.M. ; Oost, Lynette J. ; Kersten, Sander - \ 2018
Journal of Biological Chemistry 293 (2018)36. - ISSN 0021-9258 - p. 14134 - 14145.

Lipoprotein lipase (LPL) catalyzes the breakdown of circulating triglycerides in muscle and fat. LPL is inhibited by several proteins, including angiopoietin-like 4 (ANGPTL4), and may be cleaved by members of the proprotein convertase subtilisin/ kexin (PCSK) family. Here, we aimed to investigate the cleavage of LPL in adipocytes by PCSKs and study the potential involvement of ANGPTL4. A substantial portion of LPL in mouse and human adipose tissue was cleaved into N- and C-terminal fragments. Treatment of different adipocytes with the PCSK inhibitor decanoyl-RVKR-chloromethyl ketone markedly decreased LPL cleavage, indicating that LPL is cleaved by PCSKs. Silencing of Pcsk3/furin significantly decreased LPL cleavage in cell culture medium and lysates of 3T3-L1 adipocytes. Remarkably, PCSK-mediated cleavage of LPL in adipocytes was diminished by Angptl4 silencing and was decreased in adipocytes and adipose tissue of Angptl4/ mice. Differences in LPL cleavage between Angptl4/ and WT mice were abrogated by treatment with decanoyl-RVKR-chloromethyl ketone. Induction of ANGPTL4 in adipose tissue during fasting enhanced PCSK-mediated LPL cleavage, concurrent with decreased LPL activity, in WT but not Angptl4/ mice. In adipocytes, after removal of cell surface LPL by heparin, levels of N-terminal LPL were still markedly higher in WT compared with Angptl4/ adipocytes, suggesting that stimulation of PCSK-mediated LPL cleavage by ANGPTL4 occurs intracellularly. Finally, treating adipocytes with insulin increased full-length LPL and decreased N-terminal LPL in an ANGPTL4-dependent manner. In conclusion, ANGPTL4 promotes PCSK-mediated intracellular cleavage of LPL in adipocytes, likely contributing to regulation of LPL in adipose tissue. Our data provide further support for an intracellular action of ANGPTL4 in adipocytes.

Bowel Biofilms: Tipping Points between a Healthy and Compromised Gut?
Tytgat, Hanne L.P. ; Nobrega, Franklin L. ; Oost, John van der; Vos, Willem M. de - \ 2018
Trends in Microbiology (2018). - ISSN 0966-842X
biofilm - colorectal cancer - microbiota - tipping points

Bacterial communities are known to impact human health and disease. Mixed species biofilms, mostly pathogenic in nature, have been observed in dental and gastric infections as well as in intestinal diseases, chronic gut wounds and colon cancer. Apart from the appendix, the presence of thick polymicrobial biofilms in the healthy gut mucosa is still debated. Polymicrobial biofilms containing potential pathogens appear to be an early-warning signal of developing disease and can be regarded as a tipping point between a healthy and a diseased state of the gut mucosa. Key biofilm-forming pathogens and associated molecules hold promise as biomarkers. Criteria to distinguish microcolonies from biofilms are crucial to provide clarity when reporting biofilm-related phenomena in health and disease in the gut.

Opslibbing en vegetatie kwelder Ameland-Oost : Jaarrapportage 2017
Duin, Willem E. ; Sonneveld, Cor - \ 2018
Den Helder : Wageningen Marine Research (Wageningen Marine Research rapport C042/18) - 50
Twee belangrijke wetenschapsprijzen uitgereikt
Oost, John van der - \ 2018
Development of a transcriptional regulator-based bioreporter : towards a generic selection method for novel enzymes
Rossum, Teunke van - \ 2018
Wageningen University. Promotor(en): John van der Oost, co-promotor(en): Servé Kengen. - Wageningen : Wageningen University - ISBN 9789463433266 - 240
Access denied : A closer look at anti-phage defense mechanisms
Vlot, Marnix - \ 2018
Wageningen University. Promotor(en): John van der Oost, co-promotor(en): Stan Brouns. - Wageningen : Wageningen University - ISBN 9789463437752 - 178
Engineering Geobacillus thermodenitrificans to introduce cellulolytic activity; expression of native and heterologous cellulase genes
Daas, Martinus J.A. ; Nijsse, Bart ; Weijer, Antonius H.P. van de; Groenendaal, Bart W.A.J. ; Janssen, Fons ; Oost, John van der; Kranenburg, Richard van - \ 2018
BMC Biotechnology 18 (2018)1. - ISSN 1472-6750
CBP - Cellulase - Geobacillus - Metagenome - β-Xylosidase

Background: Consolidated bioprocessing (CBP) is a cost-effective approach for the conversion of lignocellulosic biomass to biofuels and biochemicals. The enzymatic conversion of cellulose to glucose requires the synergistic action of three types of enzymes: exoglucanases, endoglucanases and β-glucosidases. The thermophilic, hemicellulolytic Geobacillus thermodenitrificans T12 was shown to harbor desired features for CBP, although it lacks the desired endo and exoglucanases required for the conversion of cellulose. Here, we report the expression of both endoglucanase and exoglucanase encoding genes by G. thermodenitrificans T12, in an initial attempt to express cellulolytic enzymes that complement the enzymatic machinery of this strain. Results: A metagenome screen was performed on 73 G. thermodenitrificans strains using HMM profiles of all known CAZy families that contain endo and/or exoglucanases. Two putative endoglucanases, GE39 and GE40, belonging to glucoside hydrolase family 5 (GH5) were isolated and expressed in both E. coli and G. thermodenitrificans T12. Structure modeling of GE39 revealed a folding similar to a GH5 exo-1,3-β-glucanase from S. cerevisiae. However, we determined GE39 to be a β-xylosidase having pronounced activity towards p-nitrophenyl-β-d-xylopyranoside. Structure modelling of GE40 revealed its protein architecture to be similar to a GH5 endoglucanase from B. halodurans, and its endoglucanase activity was confirmed by enzymatic activity against 2-hydroxyethylcellulose, carboxymethylcellulose and barley β-glucan. Additionally, we introduced expression constructs into T12 containing Geobacillus sp. 70PC53 endoglucanase gene celA and both endoglucanase genes (M1 and M2) from Geobacillus sp. WSUCF1. Finally, we introduced expression constructs into T12 containing the C. thermocellum exoglucanases celK and celS genes and the endoglucanase celC gene. Conclusions: We identified a novel G. thermodenitrificans β-xylosidase (GE39) and a novel endoglucanase (GE40) using a metagenome screen based on multiple HMM profiles. We successfully expressed both genes in E. coli and functionally expressed the GE40 endoglucanase in G. thermodenitrificans T12. Additionally, the heterologous production of active CelK, a C. thermocellum derived exoglucanase, and CelA, a Geobacillus derived endoglucanase, was demonstrated with strain T12. The native hemicellulolytic activity and the heterologous cellulolytic activity described in this research provide a good basis for the further development of G. thermodenitrificans T12 as a host for consolidated bioprocessing.

Shooting the messenger : RNA-targetting CRISPR-Cas systems
Zhu, Yifan ; Klompe, Sanne E. ; Vlot, Marnix ; Oost, John van der; Staals, Raymond H.J. - \ 2018
Bioscience Reports 38 (2018)3. - ISSN 0144-8463

Since the discovery of CRISPR-Cas (Clustered Regularly Interspaced Short Palindromic Repeats, CRISPR-associated genes) immune systems, astonishing progress has been made on revealing their mechanistic foundations. Due to the immense potential as genome engineering tools, research has mainly focussed on a subset of Cas nucleases that target DNA. In addition, however, distinct types of RNA-targetting CRISPR-Cas systems have been identified. The focus of this review will be on the interference mechanisms of the RNA targetting type III and type VI CRISPR-Cas systems, their biological relevance and their potential for applications.

Genome editing by natural and engineered CRISPR-associated nucleases
Wu, Wen Y. ; Lebbink, Joyce H.G. ; Kanaar, Roland ; Geijsen, Niels ; Oost, John van der - \ 2018
Nature Chemical Biology 14 (2018)7. - ISSN 1552-4450 - p. 642 - 651.

Over the last decade, research on distinct types of CRISPR systems has revealed many structural and functional variations. Recently, several novel types of single-polypeptide CRISPR-associated systems have been discovered including Cas12a/Cpf1 and Cas13a/C2c2. Despite distant similarities to Cas9, these additional systems have unique structural and functional features, providing new opportunities for genome editing applications. Here, relevant fundamental features of natural and engineered CRISPR-Cas variants are compared. Moreover, practical matters are discussed that are essential for dedicated genome editing applications, including nuclease regulation and delivery, target specificity, as well as host repair diversity.

Mining for novel bacterial defence systems
Barrangou, Rodolphe ; Oost, John van der - \ 2018
Nature Microbiology 3 (2018). - ISSN 2058-5276 - p. 535 - 536.
Bacteria encode many strategies to prevent or escape infection. Through the analysis of metagenomic dark matter, several novel defence systems were identified, some of which were engineered and characterized in vivo, showing that they provide resistance against viruses and plasmids.
Bacteriophage DNA glucosylation impairs target DNA binding by type I and II but not by type V CRISPR–Cas effector complexes
Vlot, Marnix ; Houkes, Joep ; Lochs, Silke J.A. ; Swarts, Daan C. ; Zheng, Peiyuan ; Kunne, Tim ; Mohanraju, Prarthana ; Anders, Carolin ; Jinek, Martin ; Oost, John Van Der; Dickman, Mark J. ; Brouns, Stan J.J. - \ 2018
Nucleic Acids Research 46 (2018)2. - ISSN 0305-1048 - p. 873 - 885.
Prokaryotes encode various host defense systems that provide protection against mobile genetic elements. Restriction–modification (R–M) and CRISPR–Cas systems mediate host defense by sequence specific targeting of invasive DNA. T-even bacteriophages employ covalent modifications of nucleobases to avoid binding and therefore cleavage of their DNA by restriction endonucleases. Here, we describe that DNA glucosylation of bacteriophage genomes affects interference of some but not all CRISPR–Cas systems. We show that glucosyl modification of 5-hydroxymethylated cytosines in the DNA of bacteriophage T4 interferes with type I-E and type II-A CRISPR–Cas systems by lowering the affinity of the Cascade and Cas9–crRNA complexes for their target DNA. On the contrary, the type V-A nuclease Cas12a (also known as Cpf1) is not impaired in binding and cleavage of glucosylated target DNA, likely due to a more open structural architecture of the protein. Our results suggest that CRISPR–Cas systems have contributed to the selective pressure on phages to develop more generic solutions to escape sequence specific host defense systems.
Converting Escherichia coli into an archaebacterium with a hybrid heterochiral membrane
Caforio, Antonella ; Siliakus, Melvin F. ; Exterkate, Marten ; Jain, Samta ; Jumde, Varsha R. ; Andringa, Ruben L.H. ; Kengen, Servé W.M. ; Minnaard, Adriaan J. ; Driessen, Arnold J.M. ; Oost, John van der - \ 2018
Proceedings of the National Academy of Sciences of the United States of America 115 (2018)14. - ISSN 0027-8424 - p. 3704 - 3709.
Archaea - Bacteria - Ether lipids - Hybrid membranes - Lipid biosynthesis
One of the main differences between bacteria and archaea concerns their membrane composition. Whereas bacterial membranes are made up of glycerol-3-phosphate ester lipids, archaeal membranes are composed of glycerol-1-phosphate ether lipids. Here, we report the construction of a stable hybrid heterochiral membrane through lipid engineering of the bacterium Escherichia coli. By boosting isoprenoid biosynthesis and heterologous expression of archaeal ether lipid biosynthesis genes, we obtained a viable E. coli strain of which the membranes contain archaeal lipids with the expected stereochemistry. It has been found that the archaeal lipid biosynthesis enzymes are relatively promiscuous with respect to their glycerol phosphate backbone and that E. coli has the unexpected potential to generate glycerol-1-phosphate. The unprecedented level of 20–30% archaeal lipids in a bacterial cell has allowed for analyzing the effect on the mixed-membrane cell’s phenotype. Interestingly, growth rates are unchanged, whereas the robustness of cells with a hybrid heterochiral membrane appeared slightly increased. The implications of these findings for evolutionary scenarios are discussed.
Progress of CRISPR-Cas based genome editing in Photosynthetic microbes
Naduthodi, M.I.S. ; Barbosa, M.J. ; Oost, J. van der - \ 2018
Biotechnology Journal 13 (2018)9. - ISSN 1860-6768
The carbon footprint caused by unsustainable development and its environmental and economic impact has become a major concern in the past few decades. Photosynthetic microbes such as microalgae and cyanobacteria are capable of accumulating value-added compounds from carbon dioxide, and have been regarded as environmentally friendly alternatives to reduce the usage of fossil fuels, thereby contributing to reducing the carbon footprint. This light-driven generation of green chemicals and biofuels has triggered the research for metabolic engineering of these photosynthetic microbes. CRISPR-Cas systems are successfully implemented across a wide range of prokaryotic and eukaryotic species for efficient genome editing. However, the inception of this genome editing tool in microalgal and cyanobacterial species took off rather slowly due to various complications. In this review, we elaborate on the established CRISPR-Cas based genome editing in various microalgal and cyanobacterial species. The complications associated with CRISPR-Cas based genome editing in these species are addressed along with possible strategies to overcome these issues. It is anticipated that in the near future this will result in improving and expanding the microalgal and cyanobacterial genome engineering toolbox.
Exploration and exploitation of the environment for novel specialized metabolites
Loureiro, Catarina ; Medema, Marnix H. ; Oost, John van der; Sipkema, Detmer - \ 2018
Current Opinion in Biotechnology 50 (2018). - ISSN 0958-1669 - p. 206 - 213.
Microorganisms are Nature's little engineers of a remarkable array of bioactive small molecules that represent most of our new drugs. The wealth of genomic and metagenomic sequence data generated in the last decade has shown that the majority of novel biosynthetic gene clusters (BGCs) is identified from cultivation-independent studies, which has led to a strong expansion of the number of microbial taxa known to harbour BGCs. The large size and repeat sequences of BGCs remain a bioinformatic challenge, but newly developed software tools have been created to overcome these issues and are paramount to identify and select the most promising BGCs for further research and exploitation. Although heterologous expression of BGCs has been the greatest challenge until now, a growing number of polyketide synthase (PKS) and non-ribosomal peptide synthetase (NRPS)-encoding gene clusters have been cloned and expressed in bacteria and fungi based on techniques that mostly rely on homologous recombination. Finally, combining ecological insights with state-of-the-art computation and molecular methodologies will allow for further comprehension and exploitation of microbial specialized metabolites.
Het winterlicht gewas : proof off principle met komkommer
Kempkes, Frank ; Meinen, Esther ; Janse, Jan ; Elings, Anne ; Visser, Pieter de; Raaphorst, Marcel ; Vukadinovic, Danijela - \ 2018
Bleiswijk : Wageningen University & Research, BU Glastuinbouw (Wageningen University & Research, BU Glastuinbouw rapport WPR-725) - 58
In de winter is licht in Nederland voor vrijwel alle gewassen de limiterende factor voor de productie. In de zoektocht naar het maximaal gebruik maken van het beperkte licht in de wintermaanden is het belangrijk het licht zo optimaal mogelijk om te zetten in gewasproductie. Om de mogelijkheden beter in kaart te brengen, is in aansluiting op de opgedane kennis in het project ‘Verbetering lichtinval winterlicht’, waar de lichttoetreding door het kasdek is geoptimaliseerd voor de wintermaanden. In dit project is de gewaskant aan bod gekomen. Het werk in dit project is gefinancierd door het onderzoeksprogramma Kas Als Energiebron. Dit onderzoeksprogramma is een samenwerkingsverband tussen het ministerie van Landbouw, Natuur en Voedselkwaliteit en LTO-Glaskracht Nederland. Voor maximalisatie van de lichtonderschepping is een oost-west pad richting de eerste stap. De raseigenschappen van het gekozen komkommer ras vragen door de variatie in bladafmeting tijdens de teelt van groot naar klein een adaptatie van het gewasmanagement aan de veranderende plantvorm in de tijd. Een smalle gootafstand om hier eenvoudig op in te spelen blijkt te veel gewasbeschadiging op te leveren. De beste resultaten kwamen naar voren bij de grootste gootafstand
Hijacking CRISPR-Cas for high-throughput bacterial metabolic engineering : advances and prospects
Mougiakos, Ioannis ; Bosma, Elleke F. ; Ganguly, Joyshree ; Oost, John van der; Kranenburg, Richard van - \ 2018
Current Opinion in Biotechnology 50 (2018). - ISSN 0958-1669 - p. 146 - 157.
High engineering efficiencies are required for industrial strain development. Due to its user-friendliness and its stringency, CRISPR-Cas-based technologies have strongly increased genome engineering efficiencies in bacteria. This has enabled more rapid metabolic engineering of both the model host Escherichia coli and non-model organisms like Clostridia, Bacilli, Streptomycetes and cyanobacteria, opening new possibilities to use these organisms as improved cell factories. The discovery of novel Cas9-like systems from diverse microbial environments will extend the repertoire of applications and broaden the range of organisms in which it can be used to create novel production hosts. This review analyses the current status of prokaryotic metabolic engineering towards the production of biotechnologically relevant products, based on the exploitation of different CRISPR-related DNA/RNA endonuclease variants.
Prokaryotic Argonaute proteins : novel genome-editing tools?
Hegge, Jorrit W. ; Swarts, Daan C. ; Oost, John van der - \ 2018
Nature Reviews Microbiology 16 (2018)1. - ISSN 1740-1526 - p. 5 - 11.

Argonaute proteins constitute a highly diverse family of nucleic acid-guided proteins. They were first discovered in eukaryotes as key proteins in RNA interference systems, but homologous prokaryotic Argonaute proteins (pAgos) have also been found in archaea and bacteria. In this Progress article, we focus on long pAgo variants, a class of pAgos that are involved in nucleic acid-guided host defence against invading nucleic acids, and discuss the potential of pAgos in genome editing.

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