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“…While none of the strains grew on raw ulvan, B. licheniformis DSM13, Cryptococcus curvatus 1010 and Pseudomonas putida DSMZ 50198 consumed the monomer cocktail derived from ulvan digestion using the complete enzymatic cascade of F. agariphila that was recombinantly expressed in E. coli BL21(DE3) as described previously (Additional file 1 : Table S1, Fig. S2) [ 14 , 15 ]. Fig.…”
Section: Resultsmentioning
confidence: 99%
“…S4). These enzymatically digested ulvan-extracts cover different levels of ulvan depolymerisation as described in our previous studies and thus differ in their mono- and oligosaccharide composition [ 14 , 15 ]. Again, each hydrolysate, including the aforementioned UHB, was produced using selected F. agariphila ulvan-degrading enzymes recombinantly expressed in E. coli BL21(DE3) (Additional file 1 : Table S1, Fig.…”
Section: Resultsmentioning
confidence: 99%
“…These so-called polysaccharide utilization loci (PUL) encode proteins to mediate binding, degradation and uptake of saccharides [ 13 ]. Recently, we were able to elucidate a complex enzymatic cascade to completely deconstruct polymeric ulvan to monomeric sugar compounds using enzymes from the marine flavobacterium Formosa agariphila KM3901 T recombinantly expressed in Escherichia coli [ 14 , 15 ]. Ulvan is the main cell wall polysaccharide in the green seaweed Ulva spp.…”
Background
Marine algae are responsible for half of the global primary production, converting carbon dioxide into organic compounds like carbohydrates. Particularly in eutrophic waters, they can grow into massive algal blooms. This polysaccharide rich biomass represents a cheap and abundant renewable carbon source. In nature, the diverse group of polysaccharides is decomposed by highly specialized microbial catabolic systems. We elucidated the complete degradation pathway of the green algae-specific polysaccharide ulvan in previous studies using a toolbox of enzymes discovered in the marine flavobacterium Formosa agariphila and recombinantly expressed in Escherichia coli.
Results
In this study we show that ulvan from algal biomass can be used as feedstock for a biotechnological production strain using recombinantly expressed carbohydrate-active enzymes. We demonstrate that Bacillus licheniformis is able to grow on ulvan-derived xylose-containing oligosaccharides. Comparative growth experiments with different ulvan hydrolysates and physiological proteogenomic analyses indicated that analogues of the F. agariphila ulvan lyase and an unsaturated β-glucuronylhydrolase are missing in B. licheniformis. We reveal that the heterologous expression of these two marine enzymes in B. licheniformis enables an efficient conversion of the algal polysaccharide ulvan as carbon and energy source.
Conclusion
Our data demonstrate the physiological capability of the industrially relevant bacterium B. licheniformis to grow on ulvan. We present a metabolic engineering strategy to enable ulvan-based biorefinery processes using this bacterial cell factory. With this study, we provide a stepping stone for the development of future bioprocesses with Bacillus using the abundant marine renewable carbon source ulvan.
“…While none of the strains grew on raw ulvan, B. licheniformis DSM13, Cryptococcus curvatus 1010 and Pseudomonas putida DSMZ 50198 consumed the monomer cocktail derived from ulvan digestion using the complete enzymatic cascade of F. agariphila that was recombinantly expressed in E. coli BL21(DE3) as described previously (Additional file 1 : Table S1, Fig. S2) [ 14 , 15 ]. Fig.…”
Section: Resultsmentioning
confidence: 99%
“…S4). These enzymatically digested ulvan-extracts cover different levels of ulvan depolymerisation as described in our previous studies and thus differ in their mono- and oligosaccharide composition [ 14 , 15 ]. Again, each hydrolysate, including the aforementioned UHB, was produced using selected F. agariphila ulvan-degrading enzymes recombinantly expressed in E. coli BL21(DE3) (Additional file 1 : Table S1, Fig.…”
Section: Resultsmentioning
confidence: 99%
“…These so-called polysaccharide utilization loci (PUL) encode proteins to mediate binding, degradation and uptake of saccharides [ 13 ]. Recently, we were able to elucidate a complex enzymatic cascade to completely deconstruct polymeric ulvan to monomeric sugar compounds using enzymes from the marine flavobacterium Formosa agariphila KM3901 T recombinantly expressed in Escherichia coli [ 14 , 15 ]. Ulvan is the main cell wall polysaccharide in the green seaweed Ulva spp.…”
Background
Marine algae are responsible for half of the global primary production, converting carbon dioxide into organic compounds like carbohydrates. Particularly in eutrophic waters, they can grow into massive algal blooms. This polysaccharide rich biomass represents a cheap and abundant renewable carbon source. In nature, the diverse group of polysaccharides is decomposed by highly specialized microbial catabolic systems. We elucidated the complete degradation pathway of the green algae-specific polysaccharide ulvan in previous studies using a toolbox of enzymes discovered in the marine flavobacterium Formosa agariphila and recombinantly expressed in Escherichia coli.
Results
In this study we show that ulvan from algal biomass can be used as feedstock for a biotechnological production strain using recombinantly expressed carbohydrate-active enzymes. We demonstrate that Bacillus licheniformis is able to grow on ulvan-derived xylose-containing oligosaccharides. Comparative growth experiments with different ulvan hydrolysates and physiological proteogenomic analyses indicated that analogues of the F. agariphila ulvan lyase and an unsaturated β-glucuronylhydrolase are missing in B. licheniformis. We reveal that the heterologous expression of these two marine enzymes in B. licheniformis enables an efficient conversion of the algal polysaccharide ulvan as carbon and energy source.
Conclusion
Our data demonstrate the physiological capability of the industrially relevant bacterium B. licheniformis to grow on ulvan. We present a metabolic engineering strategy to enable ulvan-based biorefinery processes using this bacterial cell factory. With this study, we provide a stepping stone for the development of future bioprocesses with Bacillus using the abundant marine renewable carbon source ulvan.
“…The dehydratase‐catalyzed formation of an endocyclic carbon‐carbon bond in 3’‐deoxy‐3’,4’‐didehydro‐uridine‐5’‐aldehyde, an interesting structural component of uridyl peptide antibiotics, has been demonstrated by the Pac13‐catalyzed dehydration of uridine‐5’‐aldehyde [93] . The oligosaccharide dehydratase P29_PDnc has been discovered, which catalyzes the dehydration of ulvan oligosaccharides, whereby 5‐dehydro‐4‐deoxy‐ d ‐glucuronate is formed by elimination water from β‐ d ‐glucuronic acid, which is 1,4‐linked to α‐ l ‐rhamnose‐3‐sulfate [94] …”
“…[93] The oligosaccharide dehydratase P29_PDnc has been discovered, which catalyzes the dehydration of ulvan oligosaccharides, whereby 5-dehydro-4deoxy-d-glucuronate is formed by elimination water from β-dglucuronic acid, which is 1,4-linked to α-l-rhamnose-3-sulfate. [94] Aldoxime dehydratases are of much interest for the biocatalytic conversion (see Scheme 6) of aldoximes to aliphatic and aromatic nitriles. [95][96][97] Efficient conversions of aromatic aldoximes to the corresponding nitriles at high substrate concentrations using recombinant aldoxime dehydratase OxdB from Bacillus sp.…”
Biobased raw materials, such as carbohydrates, amino acids, nucleotides, or lipids contain valuable functional groups with oxygen and nitrogen atoms. An abundance of many functional groups of the same type, such as primary or secondary hydroxy groups in carbohydrates, however, limits the synthetic usefulness if similar reactivities cannot be differentiated. Therefore, selective defunctionalization of highly functionalized biobased starting materials to differentially functionalized compounds can provide a sustainable access to chiral synthons, even in case of products with fewer functional groups. Selective defunctionalization reactions, without affecting other functional groups of the same type, are of fundamental interest for biocatalytic reactions. Controlled biocatalytic defunctionalizations of biobased raw materials are attractive for obtaining valuable platform chemicals and building blocks. The biocatalytic removal of functional groups, an important feature of natural metabolic pathways, can also be utilized in a systemic strategy for sustainable metabolite synthesis.
Oceans cover 71 % of Earth's surface and are home to hundreds of thousands of species, many of which are microbial. Knowledge about marine microbes has strongly increased in the past decades due to global sampling expeditions, and hundreds of detailed studies on marine microbial ecology, physiology, and biogeochemistry. However, the translation of this knowledge into biotechnological applications or synthetic biology approaches using marine microbes has been limited so far. This review highlights key examples of marine bacteria in synthetic biology and metabolic engineering, and outlines possible future work based on the emerging marine chassis organisms Vibrio natriegens and Halomonas bluephagenesis. Furthermore, the valorization of algal polysaccharides by genetically enhanced microbes is presented as an example of the opportunities and challenges associated with blue biotechnology. Finally, new roles for marine synthetic biology in tackling pressing global challenges, including climate change and marine pollution, are discussed.
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