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1.
Environ Microbiol ; 25(12): 3192-3206, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-37722696

RESUMO

Coastal marine habitats constitute hotspots of primary productivity. In temperate regions, this is due both to massive phytoplankton blooms and dense colonisation by macroalgae that mostly store carbon as glycans, contributing substantially to local and global carbon sequestration. Because they control carbon and energy fluxes, algae-degrading microorganisms are crucial for coastal ecosystem functions. Environmental surveys revealed consistent seasonal dynamics of alga-associated bacterial assemblages, yet resolving what factors regulate the in situ abundance, growth rate and ecological functions of individual taxa remains a challenge. Here, we specifically investigated the seasonal dynamics of abundance and activity for a well-known alga-degrading marine flavobacterial genus in a tidally mixed coastal habitat of the Western English Channel. We show that members of the genus Zobellia are a stable, low-abundance component of healthy macroalgal microbiota and can also colonise particles in the water column. This genus undergoes recurring seasonal variations with higher abundances in winter, significantly associated to biotic and abiotic variables. Zobellia can become a dominant part of bacterial communities on decaying macroalgae, showing a strong activity and high estimated in situ growth rates. These results provide insights into the seasonal dynamics and environmental constraints driving natural populations of alga-degrading bacteria that influence coastal carbon cycling.


Assuntos
Flavobacteriaceae , Microbiota , Ecossistema , Estações do Ano , Carbono , Polissacarídeos
2.
Artigo em Inglês | MEDLINE | ID: mdl-37266991

RESUMO

Strain LLG6346-3.1T, isolated from the thallus of the brown alga Ericaria zosteroides collected from the Mediterranean Sea near Bastia in Corsica, France, was characterised using a polyphasic method. Cells were Gram-stain-negative, strictly aerobic, non-flagellated, motile by gliding, rod-shaped and grew optimally at 30-33 °C, at pH 8-8.5 and with 4-5 % NaCl. LLG6346-3.1T used the seaweed polysaccharide alginic acid as a sole carbon source which was vigorously liquefied. The results of phylogenetic analyses indicated that the bacterium is affiliated to the genus Zobellia (family Flavobacteriaceae, class Flavobacteriia). LLG6346-3.1T exhibited 16S rRNA gene sequence similarity values of 98.6 and 98.3 % to the type strains of Zobellia russellii and Zobellia roscoffensis, respectively, and of 97.4-98.5 % to members of other species of the genus Zobellia. The DNA G+C content of LLG6346-3.1T was determined to be 38.3 mol%. Digital DNA-DNA hybridisation predictions by the average nucleotide identity (ANI) and genome to genome distance calculator (GGDC) methods between LLG6346-3.1T and other members of the genus Zobellia showed values of 76-88 % and below 37 %, respectively. The results of phenotypic, phylogenetic and genomic analyses indicate that LLG6346-3.1T is distinct from species of the genus Zobellia with validly published names and that it represents a novel species of the genus Zobellia, for which the name Zobellia alginiliquefaciens sp. nov. is proposed. The type strain is LLG6346-3.1T (= RCC7657T = LMG 32918T).


Assuntos
Flavobacteriaceae , Phaeophyceae , Flavobacterium/genética , Ácidos Graxos/química , Filogenia , RNA Ribossômico 16S/genética , DNA Bacteriano/genética , Análise de Sequência de DNA , Composição de Bases , Técnicas de Tipagem Bacteriana , Água do Mar/microbiologia
3.
Semin Cell Dev Biol ; 134: 112-124, 2023 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-35307283

RESUMO

In brown algae, the extracellular matrix (ECM) and its constitutive polymers play crucial roles in specialized functions, including algal growth and development. In this review we offer an integrative view of ECM construction in brown algae. We briefly report the chemical composition of its main constituents, and how these are interlinked in a structural model. We examine the ECM assembly at the tissue and cell level, with consideration on its structure in vivo and on the putative subcellular sites for the synthesis of its main constituents. We further discuss the biosynthetic pathways of two major polysaccharides, alginates and sulfated fucans, and the progress made beyond the candidate genes with the biochemical validation of encoded proteins. Key enzymes involved in the elongation of the glycan chains are still unknown and predictions have been made at the gene level. Here, we offer a re-examination of some glycosyltransferases and sulfotransferases from published genomes. Overall, our analysis suggests novel investigations to be performed at both the cellular and biochemical levels. First, to depict the location of polysaccharide structures in tissues. Secondly, to identify putative actors in the ECM synthesis to be functionally studied in the future.


Assuntos
Phaeophyceae , Phaeophyceae/genética , Phaeophyceae/química , Phaeophyceae/metabolismo , Polissacarídeos/química , Polissacarídeos/metabolismo , Genoma , Matriz Extracelular/metabolismo
4.
Nucleic Acids Res ; 51(D1): D647-D653, 2023 01 06.
Artigo em Inglês | MEDLINE | ID: mdl-36318251

RESUMO

SulfAtlas (https://sulfatlas.sb-roscoff.fr/) is a knowledge-based resource dedicated to a sequence-based classification of sulfatases. Currently four sulfatase families exist (S1-S4) and the largest family (S1, formylglycine-dependent sulfatases) is divided into subfamilies by a phylogenetic approach, each subfamily corresponding to either a single characterized specificity (or few specificities in some cases) or to unknown substrates. Sequences are linked to their biochemical and structural information according to an expert scrutiny of the available literature. Database browsing was initially made possible both through a keyword search engine and a specific sequence similarity (BLAST) server. In this article, we will briefly summarize the experimental progresses in the sulfatase field in the last 6 years. To improve and speed up the (sub)family assignment of sulfatases in (meta)genomic data, we have developed a new, freely-accessible search engine using Hidden Markov model (HMM) for each (sub)family. This new tool (SulfAtlas HMM) is also a key part of the internal pipeline used to regularly update the database. SulfAtlas resource has indeed significantly grown since its creation in 2016, from 4550 sequences to 162 430 sequences in August 2022.


Assuntos
Sulfatases , Humanos , Filogenia , Sulfatases/genética , Sulfatases/química , Bases de Dados Factuais
6.
Nat Chem Biol ; 18(8): 841-849, 2022 08.
Artigo em Inglês | MEDLINE | ID: mdl-35710619

RESUMO

Sulfated glycans are ubiquitous nutrient sources for microbial communities that have coevolved with eukaryotic hosts. Bacteria metabolize sulfated glycans by deploying carbohydrate sulfatases that remove sulfate esters. Despite the biological importance of sulfatases, the mechanisms underlying their ability to recognize their glycan substrate remain poorly understood. Here, we use structural biology to determine how sulfatases from the human gut microbiota recognize sulfated glycans. We reveal seven new carbohydrate sulfatase structures spanning four S1 sulfatase subfamilies. Structures of S1_16 and S1_46 represent novel structures of these subfamilies. Structures of S1_11 and S1_15 demonstrate how non-conserved regions of the protein drive specificity toward related but distinct glycan targets. Collectively, these data reveal that carbohydrate sulfatases are highly selective for the glycan component of their substrate. These data provide new approaches for probing sulfated glycan metabolism while revealing the roles carbohydrate sulfatases play in host glycan catabolism.


Assuntos
Microbioma Gastrointestinal , Sulfatases , Bactérias/metabolismo , Humanos , Polissacarídeos/química , Sulfatases/química , Sulfatos/química
7.
ISME J ; 16(8): 2027-2039, 2022 08.
Artigo em Inglês | MEDLINE | ID: mdl-35589967

RESUMO

Macroalgae represent huge amounts of biomass worldwide, largely recycled by marine heterotrophic bacteria. We investigated the strategies of bacteria within the flavobacterial genus Zobellia to initiate the degradation of whole algal tissues, which has received little attention compared to the degradation of isolated polysaccharides. Zobellia galactanivorans DsijT has the capacity to use fresh brown macroalgae as a sole carbon source and extensively degrades algal tissues via the secretion of extracellular enzymes, even in the absence of physical contact with the algae. Co-cultures experiments with the non-degrading strain Tenacibaculum aestuarii SMK-4T showed that Z. galactanivorans can act as a pioneer that initiates algal breakdown and shares public goods with other bacteria. A comparison of eight Zobellia strains, and strong transcriptomic shifts in Z. galactanivorans cells using fresh macroalgae vs. isolated polysaccharides, revealed potential overlooked traits of pioneer bacteria. Besides brown algal polysaccharide degradation, they notably include oxidative stress resistance proteins, type IX secretion system proteins and novel uncharacterized polysaccharide utilization loci. Overall, this work highlights the relevance of studying fresh macroalga degradation to fully understand the metabolic and ecological strategies of pioneer microbial degraders, key players in macroalgal biomass remineralization.


Assuntos
Alga Marinha , Metabolismo dos Carboidratos , Polissacarídeos/metabolismo
8.
Syst Appl Microbiol ; 44(6): 126269, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34673434

RESUMO

The flavobacterial genus Zobellia is considered as a model to study macroalgal polysaccharide degradation. The lack of data regarding its prevalence and abundance in coastal habitats constitutes a bottleneck to assess its ecological strategies. To overcome this issue, real-time quantitative PCR (qPCR) and fluorescence in situ hybridization (FISH) methods targeting the 16S rRNA gene were optimized to specifically detect and quantify Zobellia on the surface of diverse macroalgae. The newly designed qPCR primers and FISH probes targeted 98 and 100% of the Zobellia strains in silico and their specificity was confirmed using pure bacterial cultures. The dynamic range of the qPCR assay spanned 8 orders of magnitude from 10 to 108 16S rRNA gene copies and the detection limit was 0.01% relative abundance of Zobellia in environmental samples. Zobellia-16S rRNA gene copies were detected on all surveyed brown, green and red macroalgae, in proportion varying between 0.1 and 0.9% of the total bacterial copies. The absolute and relative abundance of Zobellia varied with tissue aging on the kelp Laminaria digitata. Zobellia cells were successfully visualized in Ulva lactuca and stranded Palmaria palmata surface biofilm using CARD-FISH, representing in the latter 105Zobellia cells·cm-2 and 0.43% of total bacterial cells. Overall, qPCR and CARD-FISH assays enabled robust detection, quantification and localization of Zobellia representatives in complex samples, underlining their ecological relevance as primary biomass degraders potentially cross-feeding other microorganisms.


Assuntos
Flavobacteriaceae , Alga Marinha , Flavobacteriaceae/genética , Hibridização in Situ Fluorescente , RNA Ribossômico 16S/genética , Água do Mar
9.
Nature ; 598(7880): 332-337, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34616040

RESUMO

Humans have co-evolved with a dense community of microbial symbionts that inhabit the lower intestine. In the colon, secreted mucus creates a barrier that separates these microorganisms from the intestinal epithelium1. Some gut bacteria are able to utilize mucin glycoproteins, the main mucus component, as a nutrient source. However, it remains unclear which bacterial enzymes initiate degradation of the complex O-glycans found in mucins. In the distal colon, these glycans are heavily sulfated, but specific sulfatases that are active on colonic mucins have not been identified. Here we show that sulfatases are essential to the utilization of distal colonic mucin O-glycans by the human gut symbiont Bacteroides thetaiotaomicron. We characterized the activity of 12 different sulfatases produced by this species, showing that they are collectively active on all known sulfate linkages in O-glycans. Crystal structures of three enzymes provide mechanistic insight into the molecular basis of substrate specificity. Unexpectedly, we found that a single sulfatase is essential for utilization of sulfated O-glycans in vitro and also has a major role in vivo. Our results provide insight into the mechanisms of mucin degradation by a prominent group of gut bacteria, an important process for both normal microbial gut colonization2 and diseases such as inflammatory bowel disease3.


Assuntos
Bacteroides/enzimologia , Colo/metabolismo , Colo/microbiologia , Microbioma Gastrointestinal , Mucinas/metabolismo , Sulfatases/metabolismo , Acetilgalactosamina/química , Acetilgalactosamina/metabolismo , Animais , Colo/química , Cristalografia por Raios X , Feminino , Galactose/metabolismo , Humanos , Masculino , Camundongos , Modelos Moleculares , Especificidade por Substrato , Sulfatases/química
11.
Artigo em Inglês | MEDLINE | ID: mdl-34346862

RESUMO

Four marine bacterial strains were isolated from a thallus of the brown alga Ascophyllum nodosum collected in Roscoff, France. Cells were Gram-stain-negative, strictly aerobic, non-flagellated, gliding, rod-shaped and grew optimally at 25-30 °C, at pH 7-8 and with 2-4 % NaCl. Phylogenetic analyses of their 16S rRNA gene sequences showed that the bacteria were affiliated to the genus Zobellia (family Flavobacteriaceae, phylum Bacteroidetes). The four strains exhibited 97.8-100 % 16S rRNA gene sequence similarity values among themselves, 97.9-99.1 % to the type strains of Zobellia amurskyensis KMM 3526T and Zobellia laminariae KMM 3676T, and less than 99 % to other species of the genus Zobellia. The DNA G+C content of the four strains ranged from 36.7 to 37.7 mol%. Average nucleotide identity and digital DNA-DNA hybridization calculations between the new strains and other members of the genus Zobellia resulted in values of 76.4-88.9 % and below 38.5 %, respectively. Phenotypic, phylogenetic and genomic analyses showed that the four strains are distinct from species of the genus Zobellia with validly published names. They represent two novel species of the genus Zobellia, for which the names Zobellia roscoffensis sp. nov. and Zobellia nedashkovskayae sp. nov. are proposed with Asnod1-F08T (RCC6906T=KMM 6823T=CIP 111902T) and Asnod2-B07-BT (RCC6908T=KMM 6825T=CIP 111904T), respectively, as the type strains.


Assuntos
Ascophyllum , Flavobacteriaceae , Filogenia , Ascophyllum/microbiologia , Técnicas de Tipagem Bacteriana , Composição de Bases , DNA Bacteriano/genética , Ácidos Graxos/química , Flavobacteriaceae/classificação , Flavobacteriaceae/isolamento & purificação , França , Microbiota , Hibridização de Ácido Nucleico , RNA Ribossômico 16S/genética , Água do Mar , Análise de Sequência de DNA
12.
BMC Biol ; 19(1): 1, 2021 01 06.
Artigo em Inglês | MEDLINE | ID: mdl-33407428

RESUMO

BACKGROUND: Dinoflagellates are aquatic protists particularly widespread in the oceans worldwide. Some are responsible for toxic blooms while others live in symbiotic relationships, either as mutualistic symbionts in corals or as parasites infecting other protists and animals. Dinoflagellates harbor atypically large genomes (~ 3 to 250 Gb), with gene organization and gene expression patterns very different from closely related apicomplexan parasites. Here we sequenced and analyzed the genomes of two early-diverging and co-occurring parasitic dinoflagellate Amoebophrya strains, to shed light on the emergence of such atypical genomic features, dinoflagellate evolution, and host specialization. RESULTS: We sequenced, assembled, and annotated high-quality genomes for two Amoebophrya strains (A25 and A120), using a combination of Illumina paired-end short-read and Oxford Nanopore Technology (ONT) MinION long-read sequencing approaches. We found a small number of transposable elements, along with short introns and intergenic regions, and a limited number of gene families, together contribute to the compactness of the Amoebophrya genomes, a feature potentially linked with parasitism. While the majority of Amoebophrya proteins (63.7% of A25 and 59.3% of A120) had no functional assignment, we found many orthologs shared with Dinophyceae. Our analyses revealed a strong tendency for genes encoded by unidirectional clusters and high levels of synteny conservation between the two genomes despite low interspecific protein sequence similarity, suggesting rapid protein evolution. Most strikingly, we identified a large portion of non-canonical introns, including repeated introns, displaying a broad variability of associated splicing motifs never observed among eukaryotes. Those introner elements appear to have the capacity to spread over their respective genomes in a manner similar to transposable elements. Finally, we confirmed the reduction of organelles observed in Amoebophrya spp., i.e., loss of the plastid, potential loss of a mitochondrial genome and functions. CONCLUSION: These results expand the range of atypical genome features found in basal dinoflagellates and raise questions regarding speciation and the evolutionary mechanisms at play while parastitism was selected for in this particular unicellular lineage.


Assuntos
Evolução Biológica , DNA de Protozoário/análise , Dinoflagellida/citologia , Dinoflagellida/genética , Organelas/fisiologia , Proteínas de Protozoários/análise , Sequência de Bases , Evolução Molecular , Íntrons/fisiologia
13.
Int J Syst Evol Microbiol ; 69(8): 2514-2521, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31199221

RESUMO

Strain 1T, isolated in the 1970s from the thallus of the carrageenophytic red algae, Eucheuma spinosum, collected in Hawaii, USA, was characterized using a polyphasic method. Cells were Gram-stain-negative, strictly aerobic, non-flagellated, ovoid or rod-shaped and grew optimally at 20-25 °C, at pH 6-9 and with 2-4 % NaCl. Strain 1T used the seaweed polysaccharides ι-carrageenan, laminarin and alginic acid as sole carbon sources. The major fatty acids were C16 : 0, C18 : 1 ω7c and summed feature 3 (C16 : 1 ω7c and/or iso-C15 : 0 2OH) with significant amounts (>6 %) of C16 : 0 N alcohol and 10 methyl C17 : 0. The respiratory quinone was Q-8 and major polar lipids were phosphatidylethanolamine, phosphatidylglycerol and an unknown aminolipid. Phylogenetic analyses showed that the bacterium is affiliated to the genus Alteromonas (family Alteromonadaceae, class Gammaproteobacteria). Strain 1T exhibited 16S rRNA gene sequence similarity values of 98.8 and 99.2 % to the type strains of Alteromonas mediterranea and Alteromonas australica respectively, and of 95.2-98.6 % to other species of the genus Alteromonas. The DNA G+C content of strain 1T was determined to be 43.9 mol%. Digital DNA-DNA hybridization predictions by the ANI and GGDC methods between strain 1T and other members of the genus Alteromonas showed values below 83 % and 30 %, respectively. The phenotypic, phylogenetic and genomic analyses show that strain 1T is distinct from species of the genus Alteromonas with validly published names and that it represents a novel species of the genus Alteromonas, for which the name Alteromonasfortis sp. nov. is proposed. The type strain is 1T (=ATCC 43554T=RCC 5933T=CIP 111645T=DSM 106819T).


Assuntos
Alteromonas/classificação , Carragenina/metabolismo , Filogenia , Rodófitas/microbiologia , Alteromonas/isolamento & purificação , Técnicas de Tipagem Bacteriana , Composição de Bases , DNA Bacteriano/genética , Ácidos Graxos/química , Havaí , Hibridização de Ácido Nucleico , Fosfatidiletanolaminas/química , Fosfatidilgliceróis/química , RNA Ribossômico 16S/genética , Alga Marinha/microbiologia , Análise de Sequência de DNA
14.
Front Microbiol ; 9: 2740, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30524390

RESUMO

About half of seaweed biomass is composed of polysaccharides. Most of these complex polymers have a marked polyanionic character. For instance, the red algal cell wall is mainly composed of sulfated galactans, agars and carrageenans, while brown algae contain alginate and fucose-containing sulfated polysaccharides (FCSP) as cell wall polysaccharides. Some marine heterotrophic bacteria have developed abilities to grow on such macroalgal polysaccharides. This is the case of Pseudoalteromonas carrageenovora 9T (ATCC 43555T), a marine gammaproteobacterium isolated in 1955 and which was an early model organism for studying carrageenan catabolism. We present here the genomic analysis of P. carrageenovora. Its genome is composed of two chromosomes and of a large plasmid encompassing 109 protein-coding genes. P. carrageenovora possesses a diverse repertoire of carbohydrate-active enzymes (CAZymes), notably specific for the degradation of macroalgal polysaccharides (laminarin, alginate, FCSP, carrageenans). We confirm these predicted capacities by screening the growth of P. carrageenovora with a large collection of carbohydrates. Most of these CAZyme genes constitute clusters located either in the large chromosome or in the small one. Unexpectedly, all the carrageenan catabolism-related genes are found in the plasmid, suggesting that P. carrageenovora acquired its hallmark capacity for carrageenan degradation by horizontal gene transfer (HGT). Whereas P. carrageenovora is able to use lambda-carrageenan as a sole carbon source, genomic and physiological analyses demonstrate that its catabolic pathway for kappa- and iota-carrageenan is incomplete. This is due to the absence of the recently discovered 3,6-anhydro-D-galactosidase genes (GH127 and GH129 families). A genomic comparison with 52 Pseudoalteromonas strains confirms that carrageenan catabolism has been recently acquired only in a few species. Even though the loci for cellulose biosynthesis and alginate utilization are located on the chromosomes, they were also horizontally acquired. However, these HGTs occurred earlier in the evolution of the Pseudoalteromonas genus, the cellulose- and alginate-related loci being essentially present in one large, late-diverging clade (LDC). Altogether, the capacities to degrade cell wall polysaccharides from macroalgae are not ancestral in the Pseudoalteromonas genus. Such catabolism in P. carrageenovora resulted from a succession of HGTs, likely allowing an adaptation to the life on the macroalgal surface.

15.
ISME J ; 12(12): 2894-2906, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30061707

RESUMO

Polysaccharide degradation by heterotrophic microbes is a key process within Earth's carbon cycle. Here, we use environmental proteomics and metagenomics in combination with cultivation experiments and biochemical characterizations to investigate the molecular details of in situ polysaccharide degradation mechanisms during microalgal blooms. For this, we use laminarin as a model polysaccharide. Laminarin is a ubiquitous marine storage polymer of marine microalgae and is particularly abundant during phytoplankton blooms. In this study, we show that highly specialized bacterial strains of the Bacteroidetes phylum repeatedly reached high abundances during North Sea algal blooms and dominated laminarin turnover. These genomically streamlined bacteria of the genus Formosa have an expanded set of laminarin hydrolases and transporters that belonged to the most abundant proteins in the environmental samples. In vitro experiments with cultured isolates allowed us to determine the functions of in situ expressed key enzymes and to confirm their role in laminarin utilization. It is shown that laminarin consumption of Formosa spp. is paralleled by enhanced uptake of diatom-derived peptides. This study reveals that genome reduction, enzyme fusions, transporters, and enzyme expansion as well as a tight coupling of carbon and nitrogen metabolism provide the tools, which make Formosa spp. so competitive during microalgal blooms.


Assuntos
Bacteroidetes/fisiologia , Eutrofização , Flavobacteriaceae/fisiologia , Glucanos/metabolismo , Microalgas/microbiologia , Polissacarídeos/metabolismo , Adaptação Fisiológica , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Bacteroidetes/genética , Ciclo do Carbono , Flavobacteriaceae/genética , Hidrolases/genética , Hidrolases/metabolismo , Metagenômica , Microalgas/metabolismo , Mar do Norte , Fitoplâncton/metabolismo , Fitoplâncton/microbiologia
16.
Front Microbiol ; 9: 839, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29774012

RESUMO

Marine microbes are a rich source of enzymes for the degradation of diverse polysaccharides. Paraglaciecola hydrolytica S66T is a marine bacterium capable of hydrolyzing polysaccharides found in the cell wall of red macroalgae. In this study, we applied an approach combining genomic mining with functional analysis to uncover the potential of this bacterium to produce enzymes for the hydrolysis of complex marine polysaccharides. A special feature of P. hydrolytica S66T is the presence of a large genomic region harboring an array of carbohydrate-active enzymes (CAZymes) notably agarases and carrageenases. Based on a first functional characterization combined with a comparative sequence analysis, we confirmed the enzymatic activities of several enzymes required for red algal polysaccharide degradation by the bacterium. In particular, we report for the first time, the discovery of novel enzyme activities targeting furcellaran, a hybrid carrageenan containing both ß-carrageenan and κ/ß-carrageenan motifs. Some of these enzymes represent a new subfamily within the CAZy classification. From the combined analyses, we propose models for the complete degradation of agar and κ/ß-type carrageenan by P. hydrolytica S66T. The novel enzymes described here may find value in new bio-based industries and advance our understanding of the mechanisms responsible for recycling of red algal polysaccharides in marine ecosystems.

17.
Nat Commun ; 8(1): 1685, 2017 11 22.
Artigo em Inglês | MEDLINE | ID: mdl-29162826

RESUMO

Macroalgae contribute substantially to primary production in coastal ecosystems. Their biomass, mainly consisting of polysaccharides, is cycled into the environment by marine heterotrophic bacteria using largely uncharacterized mechanisms. Here we describe the complete catabolic pathway for carrageenans, major cell wall polysaccharides of red macroalgae, in the marine heterotrophic bacterium Zobellia galactanivorans. Carrageenan catabolism relies on a multifaceted carrageenan-induced regulon, including a non-canonical polysaccharide utilization locus (PUL) and genes distal to the PUL, including a susCD-like pair. The carrageenan utilization system is well conserved in marine Bacteroidetes but modified in other phyla of marine heterotrophic bacteria. The core system is completed by additional functions that might be assumed by non-orthologous genes in different species. This complex genetic structure may be the result of multiple evolutionary events including gene duplications and horizontal gene transfers. These results allow for an extension on the definition of bacterial PUL-mediated polysaccharide digestion.


Assuntos
Carragenina/metabolismo , Flavobacteriaceae/genética , Flavobacteriaceae/metabolismo , Regulon , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Bacteroidetes/genética , Bacteroidetes/metabolismo , Cristalografia por Raios X , Evolução Molecular , Galactosidases/química , Galactosidases/genética , Galactosidases/metabolismo , Genes Bacterianos , Redes e Vias Metabólicas/genética , Modelos Moleculares , Família Multigênica , Filogenia , Conformação Proteica , RNA Bacteriano/genética , Análise de Sequência de RNA , Especificidade da Espécie
18.
J Biol Chem ; 292(48): 19919-19934, 2017 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-29030427

RESUMO

Carrageenans are sulfated α-1,3-ß-1,4-galactans found in the cell wall of some red algae that are practically valuable for their gelation and biomimetic properties but also serve as a potential carbon source for marine bacteria. Carbohydrate degradation has been studied extensively for terrestrial plant/bacterial systems, but sulfation is not present in these cases, meaning the marine enzymes used to degrade carrageenans must possess unique features to recognize these modifications. To gain insights into these features, we have focused on κ-carrageenases from two distant bacterial phyla, which belong to glycoside hydrolase family 16 and cleave the ß-1,4 linkage of κ-carrageenan. We have solved the crystal structure of the catalytic module of ZgCgkA from Zobellia galactanivorans at 1.66 Å resolution and compared it with the only other structure available, that of PcCgkA from Pseudoalteromonas carrageenovora 9T (ATCC 43555T). We also describe the first substrate complex in the inactivated mutant form of PcCgkA at 1.7 Å resolution. The structural and biochemical comparison of these enzymes suggests key determinants that underlie the functional properties of this subfamily. In particular, we identified several arginine residues that interact with the polyanionic substrate, and confirmed the functional relevance of these amino acids using a targeted mutagenesis strategy. These results give new insight into the diversity of the κ-carrageenase subfamily. The phylogenetic analyses show the presence of several distinct clades of enzymes that relate to differences in modes of action or subtle differences within the same substrate specificity, matching the hybrid character of the κ-carrageenan polymer.


Assuntos
Metabolismo dos Carboidratos , Flavobacteriaceae/enzimologia , Glicosídeo Hidrolases/metabolismo , Biologia Marinha , Pseudoalteromonas/enzimologia , Catálise , Cristalografia por Raios X , Glicosídeo Hidrolases/química , Glicosídeo Hidrolases/classificação , Cinética , Filogenia , Conformação Proteica , Especificidade por Substrato
19.
Front Microbiol ; 8: 1542, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28861057

RESUMO

Tenacibaculum maritimum is a devastating bacterial pathogen of wild and farmed marine fish with a broad host range and a worldwide distribution. We report here the complete genome sequence of the T. maritimum type strain NCIMB 2154T. The genome consists of a 3,435,971-base pair circular chromosome with 2,866 predicted protein-coding genes. Genes encoding the biosynthesis of exopolysaccharides, the type IX secretion system, iron uptake systems, adhesins, hemolysins, proteases, and glycoside hydrolases were identified. They are likely involved in the virulence process including immune escape, invasion, colonization, destruction of host tissues, and nutrient scavenging. Among the predicted virulence factors, type IX secretion-mediated and cell-surface exposed proteins were identified including an atypical sialidase, a sphingomyelinase and a chondroitin AC lyase which activities were demonstrated in vitro.

20.
Int J Syst Evol Microbiol ; 67(7): 2242-2247, 2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-28671532

RESUMO

A novel bacterial strain, S66T, was isolated from eelgrass collected on the coastline of Zealand, Denmark. Polyphasic analyses involving phenotypic, phylogenetic and genomic methods were used to characterize strain S66T. The strain was Gram-reaction-negative, rod-shaped, aerobic, and displayed growth at 10-25 °C (optimum 20-25 °C) and at pH 7-9 (optimum pH 7.5). Furthermore, strain S66T grew on seaweed polysaccharides agar, agarose, porphyran, κ-carrageenan, alginate and laminarin as sole carbon sources. Major fatty acids were C16 : 0, C16 : 1ω7c and C18 : 1ω7c. The respiratory quinone was determined to be Q-8, and major polar lipids were phosphatidylethanolamine and phosphatidylglycerol. The DNA G+C content was determined to be 42.2 mol%. Phylogenetic analyses based on the 16S rRNA gene and GyrB sequence comparisons showed that the bacterium was affiliated with the genus Paraglaciecola within the family Alteromonadaceae of the class Gammaproteobacteria. The percentage similarity between the 16S rRNA gene and GyrB sequences of strain S66T and other members of the genus Paraglaciecola were 94-95 % and 84-85 %, respectively. Based on the genome sequence of S66T, the average nucleotide identity (ANI) between strain S66T and other members of the genus Paraglaciecola was 77-80 %, and DNA-DNA hybridization prediction showed values of less than 24 % relatedness, respectively, between S66T and other species of the genus Paraglaciecola. The phenotypic, phylogenetic and genomic analyses support the hypothesis that strain S66T represents a novel species of the genus Paraglaciecola, for which the name Paraglaciecola hydrolytica sp. nov. is proposed. The type strain is S66T (=LMG 29457T=NCIMB 15060T=DSM 102834T).


Assuntos
Alteromonadaceae/classificação , Filogenia , Polissacarídeos/química , Alga Marinha/química , Alteromonadaceae/genética , Alteromonadaceae/isolamento & purificação , Técnicas de Tipagem Bacteriana , Composição de Bases , DNA Bacteriano/genética , Dinamarca , Ácidos Graxos/química , Genes Bacterianos , Hibridização de Ácido Nucleico , Fosfatidiletanolaminas/química , Fosfatidilgliceróis/química , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Ubiquinona/química
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