Characterization of Redox Sensitive Brown Algal Mannitol-1-Phosphatases

Macroalgae (seaweeds) are key primary producers in marine coastal habitats and largely contribute to global ocean carbon fluxes. They also represent attractive renewable feedstock for the production of biofuels, food, feed, and bioactive. Brown algae are seaweeds that produce alginates and fucose containing sulfated polysaccharides in their cell wall and laminarin and mannitol for carbon storage. The availability of genomes of the kelp Saccharina japonica and of the filamentous Ectocarpus sp. paved the way for the biochemical characterization of recombinant enzymes involved in their polysaccharide and carbohydrates synthesis, including, notably, mannitol. Brown algal mannitol biosynthesis starts with the conversion of fructose-6-phospate into mannitol-1-phosphate (mannitol-1P), and this intermediate is hydrolysed by a haloacid dehalogenase phosphatase (M1Pase) to produce mannitol. We report here the biochemical characterization of a second M1Pase in Ectocarpus sp. (EsM1Pase1). Both Ectocarpus M1Pases were redox-sensitive enzymes, with EsM1Pase1 active only in presence of the reducing agent. Such catalytic properties have not been observed for any M1Pases yet. EsM1Pases were specific to mannitol-1-P, in contrast to S. japonica M1Pases that could act on other phosphorylated sugars. Finally, brown algal M1Pases formed two well-supported clades, with possible distinct subcellular localization and physiological role(s) under diverse environmental conditions and/or life cycle stages

Study of the fungal community associated to the red macroalga Palmaria palmata for the production of cell wall degrading enzymes of biotechnological interest

La communauté fongique associée à l’algue rouge Palmaria palmata a été étudiée, en particulier pour sa production de xylanases et de cellulases, enzymes utilisées dans l’extraction de composés d’intérêt comme la R-phycoérythrine (R-PE). Dans un premier temps, des récoltes régulières de P. palmata ont été effectuées pendant un an afin de réaliser un vaste travail d’isolement et d’identification de souches fongiques. Il s’agit de la première étude globale de diversité fongique chez P. palmata. 308 souches fongiques ont été isolées, réparties dans 147 taxons appartenant majoritairement aux Ascomycètes. Ensuite, un criblage enzymatique a été réalisé permettant d’évaluer qualitativement les activités xylanolytiques et cellulolytiques de chaque champignon isolé. Il en résulte que sur les 308 souches fongiques, la majorité d’entre-elles est caractérisée par au moins l’une de ces deux activités enzymatiques. Suite à ce criblage, six souches ont été sélectionnées pour une analyse quantitative de leur double activité xylanolytique et cellulolytique.Ces souches appartiennent aux genres Cladosporium, Penicillium et Aspergillus majoritairement observés chez P. palmata. La quantification de ces activités enzymatiques a été réalisée et des tests préliminaires de dégradation de thalle de P. palmata ont été effectués afin d’extraire la R-PE. Ces tests de dégradation ont été réalisés à partir d’extraits enzymatiques bruts. Les souches MMS1733 (Aspergillus sp.), MMS1906 (Penicillium sp.) et MMS1910 (Penicillium brevicompactum) ont semblé particulièrement intéressantes pour leur production d’enzymes permettant l’extraction de R-PE. Parmi les pistes de recherches futures, la purification des enzymes issues de ces souches, leur caractérisation plus fine ainsi que leur utilisation potentielle à grande échelle pourraient aboutir à des résultats particulièrement prometteurs dans le domaine des biotechnologies marines.The fungal community associated with the red alga Palmaria palmata was studied, in particular for its production of xylanases and cellulases which are used in the extraction of compounds of interest such as the R-phycoerythrin (R-PE). Initially, over a period of one year, P. palmata was regularly harvested in order to isolate and identify strains from algal thalli. Up to now, it is the first global study dealing with fungal diversity from P. palmata. 308 fungal strains were isolated, among 147 taxa mostly belonging to the Ascomycetes. Next, an enzymatic screening was performed on fungal isolates to assess qualitatively their xylanolytic and cellulolytic activities. Among the 308 fungal strains, the majority were characterised by at least one of these two enzymatic activities. Six isolates were selected from this screening for a quantitative analysis of their double xylanolytic and cellulolytic activity.These strains belonged to the genera Cladosporium, Penicillium and Aspergillus mainly observed in P. palmata. Quantification of enzymatic activities was performed as well as preliminary tests of P. palmata thalli degradation for R-PE extraction. These degradation tests were realized by using crude enzymatic extracts. Strains MMS1733 (Aspergillus sp.), MMS1906 (Penicillium sp.) and MMS1910 (Penicillium brevicompactum) seemed particularly interesting for R-PE extraction. Among the perspectives, the purification of enzymes from these strains, their detailed characterisation and their potential large-scale use could lead to particularly promising results in the field of marine biotechnology

Etude de la communauté fongique associée à l'algue rouge Palmaria palmata pour la production d'enzymes de dégradation pariétale à visée biotechnologique

The fungal community associated with the red alga Palmaria palmata was studied, in particular for its production of xylanases and cellulases which are used in the extraction of compounds of interest such as the R-phycoerythrin (R-PE). Initially, over a period of one year, P. palmata was regularly harvested in order to isolate and identify strains from algal thalli. Up to now, it is the first global study dealing with fungal diversity from P. palmata. 308 fungal strains were isolated, among 147 taxa mostly belonging to the Ascomycetes. Next, an enzymatic screening was performed on fungal isolates to assess qualitatively their xylanolytic and cellulolytic activities. Among the 308 fungal strains, the majority were characterised by at least one of these two enzymatic activities. Six isolates were selected from this screening for a quantitative analysis of their double xylanolytic and cellulolytic activity.These strains belonged to the genera Cladosporium, Penicillium and Aspergillus mainly observed in P. palmata. Quantification of enzymatic activities was performed as well as preliminary tests of P. palmata thalli degradation for R-PE extraction. These degradation tests were realized by using crude enzymatic extracts. Strains MMS1733 (Aspergillus sp.), MMS1906 (Penicillium sp.) and MMS1910 (Penicillium brevicompactum) seemed particularly interesting for R-PE extraction. Among the perspectives, the purification of enzymes from these strains, their detailed characterisation and their potential large-scale use could lead to particularly promising results in the field of marine biotechnology.La communauté fongique associée à l’algue rouge Palmaria palmata a été étudiée, en particulier pour sa production de xylanases et de cellulases, enzymes utilisées dans l’extraction de composés d’intérêt comme la R-phycoérythrine (R-PE). Dans un premier temps, des récoltes régulières de P. palmata ont été effectuées pendant un an afin de réaliser un vaste travail d’isolement et d’identification de souches fongiques. Il s’agit de la première étude globale de diversité fongique chez P. palmata. 308 souches fongiques ont été isolées, réparties dans 147 taxons appartenant majoritairement aux Ascomycètes. Ensuite, un criblage enzymatique a été réalisé permettant d’évaluer qualitativement les activités xylanolytiques et cellulolytiques de chaque champignon isolé. Il en résulte que sur les 308 souches fongiques, la majorité d’entre-elles est caractérisée par au moins l’une de ces deux activités enzymatiques. Suite à ce criblage, six souches ont été sélectionnées pour une analyse quantitative de leur double activité xylanolytique et cellulolytique.Ces souches appartiennent aux genres Cladosporium, Penicillium et Aspergillus majoritairement observés chez P. palmata. La quantification de ces activités enzymatiques a été réalisée et des tests préliminaires de dégradation de thalle de P. palmata ont été effectués afin d’extraire la R-PE. Ces tests de dégradation ont été réalisés à partir d’extraits enzymatiques bruts. Les souches MMS1733 (Aspergillus sp.), MMS1906 (Penicillium sp.) et MMS1910 (Penicillium brevicompactum) ont semblé particulièrement intéressantes pour leur production d’enzymes permettant l’extraction de R-PE. Parmi les pistes de recherches futures, la purification des enzymes issues de ces souches, leur caractérisation plus fine ainsi que leur utilisation potentielle à grande échelle pourraient aboutir à des résultats particulièrement prometteurs dans le domaine des biotechnologies marines

Cultivable epiphytic bacteria of the Chlorophyta Ulva sp.: diversity, antibacterial, and biofilm-modulating activities

International audienceAbstract Aims Macroalgae harbor a rich epiphytic microbiota that plays a crucial role in algal morphogenesis and defense mechanisms. This study aims to isolate epiphytic cultivable microbiota from Ulva sp. surfaces. Various culture media were employed to evaluate a wide range of cultivable microbiota. Our objective was to assess the antibacterial and biofilm-modulating activities of supernatants from isolated bacteria. Methods and results Sixty-nine bacterial isolates from Ulva sp. were identified based on 16S rRNA gene sequencing. Their antibacterial activity and biofilm modulation potential were screened against three target marine bacteria: 45%, mostly affiliated with Gammaproteobacteria and mainly grown on diluted R2A medium (R2Ad), showed strong antibacterial activity, while 18% had a significant impact on biofilm modulation. Molecular network analysis was carried out on four bioactive bacterial supernatants, revealing new molecules potentially responsible for their activities. Conclusion R2Ad offered the greatest diversity and proportion of active isolates. The molecular network approach holds promise for both identifying bacterial isolates based on their molecular production and characterizing antibacterial and biofilm-modulating activities

Utilisation de biomarqueurs chez une espèce-sentinelle d'invertébré marin dans un protocole de biosurveillance de la qualité des milieux portuaires

International audienceUtilisation de biomarqueurs chez une espèce-sentinelle d'invertébré marin dans un protocole de biosurveillance de la qualité des milieux portuaire

Utilisation de biomarqueurs chez une espèce-sentinelle d'invertébré marin dans un protocole de biosurveillance de la qualité des milieux portuaires

International audienceUtilisation de biomarqueurs chez une espèce-sentinelle d'invertébré marin dans un protocole de biosurveillance de la qualité des milieux portuaire

Utilisation de biomarqueurs chez une espèce-sentinelle d'invertébré marin dans un protocole de biosurveillance de la qualité des milieux portuaires

International audienceUtilisation de biomarqueurs chez une espèce-sentinelle d'invertébré marin dans un protocole de biosurveillance de la qualité des milieux portuaire

Heterologous Expression and Biochemical Characterization of a New Chloroperoxidase Isolated from the Deep-Sea Hydrothermal Vent Black Yeast Hortaea werneckii UBOCC-A-208029

International audienceThe initiation of this study relies on a targeted genome-mining approach to highlight the presence of a putative vanadium-dependent haloperoxidase-encoding gene in the deep-sea hydrothermal vent fungus Hortaea werneckii UBOCC-A-208029. To date, only three fungal vanadium-dependent haloperoxidases have been described, one from the terrestrial species Curvularia inaequalis, one from the fungal plant pathogen Botrytis cinerea , and one from a marine derived isolate identified as Alternaria didymospora . In this study, we describe a new vanadium chloroperoxidase from the black yeast H. werneckii , successfully cloned and overexpressed in a bacterial host, which possesses higher affinity for bromide (K m = 26 µM) than chloride (K m = 237 mM). The enzyme was biochemically characterized, and we have evaluated its potential for biocatalysis by determining its stability and tolerance in organic solvents. We also describe its potential three-dimensional structure by building a model using the AlphaFold 2 artificial intelligence tool. This model shows some conservation of the 3D structure of the active site compared to the vanadium chloroperoxidase from C. inaequalis but it also highlights some differences in the active site entrance and the volume of the active site pocket, underlining its originality