scholarly journals Identification of an Oligosaccharide Dehydrogenase from Pycnoporus Cinnabarinus Provides Insights into Fungal Breakdown of Lignocellulose

2021 ◽  
Author(s):  
Gabriele Cerutti ◽  
Elena Gugole ◽  
Linda Celeste Montemiglio ◽  
Annick Turbé-Doan ◽  
Dehbia Chena ◽  
...  
Keyword(s):  
Function Analysis ◽  
Sequence Similarity ◽  
Functional Characterization ◽  
Physiological Role ◽  
Free Form ◽  
Lignocellulose Degradation ◽  
Carbohydrate Active Enzymes ◽  
Recognition Mechanism ◽  
Pycnoporus Cinnabarinus ◽  
Enzymatic Function

Abstract Background: Fungal glucose dehydrogenases (GDHs) are FAD-dependent enzymes belonging to the glucose-methanol-choline oxidoreductase superfamily. These enzymes are classified in the “Auxiliary Activity” family 3 (AA3) of the Carbohydrate-Active enZymes database, and more specifically in subfamily AA3_2, that also includes the closely related flavoenzymes aryl-alcohol oxidase and glucose 1-oxidase. Based on sequence similarity to known fungal GDHs, an AA3_2 enzyme active on glucose was identified in the genome of Pycnoporus cinnabarinus, a model Basidiomycete able to completely degrade lignin.Results: In our work, substrate screening and functional characterization showed an unexpected preferential activity of this enzyme toward oligosaccharides containing a b(1à3) glycosidic bond, with the highest efficiency observed for the disaccharide laminaribiose. Despite its sequence similarity to GDHs, we defined a novel enzymatic activity, namely oligosaccharide dehydrogenase (ODH), for this enzyme. The crystallographic structures of ODH in the sugar-free form and in complex with glucose and laminaribiose unveiled a peculiar saccharide recognition mechanism which is not shared with previously characterized AA3 oxidoreductases and accounts for ODH preferential activity toward oligosaccharides. The sugar molecules in the active site of ODH are mainly stabilized through CH-p interactions with aromatic residues rather than through hydrogen bonds with highly conserved residues, as observed instead for the fungal glucose dehydrogenases and oxidases characterized to date. Finally, three sugar-binding sites were identified on ODH external surface, which were not previously observed and might be of importance in the physiological scenario.Conclusions: Structure-function analysis of ODH is consistent with its role as an auxiliary enzyme in lignocellulose degradation and unveils yet another enzymatic function within the AA3 family of the Carbohydrate-Active enZymes database. Our findings allow deciphering the molecular determinants of substrate binding and provide insight into the physiological role of ODH, opening new perspectives to exploit biodiversity for lignocellulose transformation into fuels and chemicals.

scholarly journals Crystal structure and functional characterization of an oligosaccharide dehydrogenase from Pycnoporus cinnabarinus provides insights into fungal breakdown of lignocellulose

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Gabriele Cerutti ◽  
Elena Gugole ◽  
Linda Celeste Montemiglio ◽  
Annick Turbé-Doan ◽  
Dehbia Chena ◽  
...  
Keyword(s):  
Function Analysis ◽  
Sequence Similarity ◽  
Functional Characterization ◽  
Physiological Role ◽  
Free Form ◽  
Lignocellulose Degradation ◽  
Carbohydrate Active Enzymes ◽  
Recognition Mechanism ◽  
Pycnoporus Cinnabarinus ◽  
Enzymatic Function

Abstract Background Fungal glucose dehydrogenases (GDHs) are FAD-dependent enzymes belonging to the glucose-methanol-choline oxidoreductase superfamily. These enzymes are classified in the “Auxiliary Activity” family 3 (AA3) of the Carbohydrate-Active enZymes database, and more specifically in subfamily AA3_2, that also includes the closely related flavoenzymes aryl-alcohol oxidase and glucose 1-oxidase. Based on sequence similarity to known fungal GDHs, an AA3_2 enzyme active on glucose was identified in the genome of Pycnoporus cinnabarinus, a model Basidiomycete able to completely degrade lignin. Results In our work, substrate screening and functional characterization showed an unexpected preferential activity of this enzyme toward oligosaccharides containing a β(1→3) glycosidic bond, with the highest efficiency observed for the disaccharide laminaribiose. Despite its sequence similarity to GDHs, we defined a novel enzymatic activity, namely oligosaccharide dehydrogenase (ODH), for this enzyme. The crystallographic structures of ODH in the sugar-free form and in complex with glucose and laminaribiose unveiled a peculiar saccharide recognition mechanism which is not shared with previously characterized AA3 oxidoreductases and accounts for ODH preferential activity toward oligosaccharides. The sugar molecules in the active site of ODH are mainly stabilized through CH-π interactions with aromatic residues rather than through hydrogen bonds with highly conserved residues, as observed instead for the fungal glucose dehydrogenases and oxidases characterized to date. Finally, three sugar-binding sites were identified on ODH external surface, which were not previously observed and might be of importance in the physiological scenario. Conclusions Structure–function analysis of ODH is consistent with its role as an auxiliary enzyme in lignocellulose degradation and unveils yet another enzymatic function within the AA3 family of the Carbohydrate-Active enZymes database. Our findings allow deciphering the molecular determinants of substrate binding and provide insight into the physiological role of ODH, opening new perspectives to exploit biodiversity for lignocellulose transformation into fuels and chemicals.

scholarly journals Structural and Functional Characterization of the ABCC6 Transporter in Hepatic Cells: Role on PXE, Cancer Therapy and Drug Resistance

2021 ◽  
Vol 22 (6) ◽  
pp. 2858
Author(s):  
Faustino Bisaccia ◽  
Prashant Koshal ◽  
Vittorio Abruzzese ◽  
Maria Antonietta Castiglione Morelli ◽  
Angela Ostuni
Keyword(s):  
Cancer Therapy ◽  
Functional Characterization ◽  
Physiological Role ◽  
Pseudoxanthoma Elasticum ◽  
Connective Tissues ◽  
Functional Studies ◽  
Extracellular Milieu ◽  
Hepatic Cells ◽  
Ectopic Mineralization ◽  
Anti Cancer

Pseudoxanthoma elasticum (PXE) is a complex autosomal recessive disease caused by mutations of ABCC6 transporter and characterized by ectopic mineralization of soft connective tissues. Compared to the other ABC transporters, very few studies are available to explain the structural components and working of a full ABCC6 transporter, which may provide some idea about its physiological role in humans. Some studies suggest that mutations of ABCC6 in the liver lead to a decrease in some circulating factor and indicate that PXE is a metabolic disease. It has been reported that ABCC6 mediates the efflux of ATP, which is hydrolyzed in PPi and AMP; in the extracellular milieu, PPi gives potent anti-mineralization effect, whereas AMP is hydrolyzed to Pi and adenosine which affects some cellular properties by modulating the purinergic pathway. Structural and functional studies have demonstrated that silencing or inhibition of ABCC6 with probenecid changed the expression of several genes and proteins such as NT5E and TNAP, as well as Lamin, and CDK1, which are involved in cell motility and cell cycle. Furthermore, a change in cytoskeleton rearrangement and decreased motility of HepG2 cells makes ABCC6 a potential target for anti-cancer therapy. Collectively, these findings suggested that ABCC6 transporter performs functions that modify both the external and internal compartments of the cells.

scholarly journals Targeting spectrin redox switches to regulate the mechanoproperties of red blood cells

2020 ◽  
Vol 0 (0) ◽  
Author(s):  
Frederik Barbarino ◽  
Lucas Wäschenbach ◽  
Virginia Cavalho-Lemos ◽  
Melissa Dillenberger ◽  
Katja Becker ◽  
...  
Keyword(s):  
Mechanical Properties ◽  
Red Blood Cells ◽  
Blood Cells ◽  
Redox Regulation ◽  
Current Knowledge ◽  
Structural Characteristics ◽  
Functional Characterization ◽  
Physiological Role ◽  
Spectrin Cytoskeleton ◽  
Redox Switches

AbstractThe mechanical properties of red blood cells (RBCs) are fundamental for their physiological role as gas transporters. RBC flexibility and elasticity allow them to survive the hemodynamic changes in the different regions of the vascular tree, to dynamically contribute to the flow thereby decreasing vascular resistance, and to deform during the passage through narrower vessels. RBC mechanoproperties are conferred mainly by the structural characteristics of their cytoskeleton, which consists predominantly of a spectrin scaffold connected to the membrane via nodes of actin, ankyrin and adducin. Changes in redox state and treatment with thiol-targeting molecules decrease the deformability of RBCs and affect the structure and stability of the spectrin cytoskeleton, indicating that the spectrin cytoskeleton may contain redox switches. In this perspective review, we revise current knowledge about the structural and functional characterization of spectrin cysteine redox switches and discuss the current lines of research aiming to understand the role of redox regulation on RBC mechanical properties. These studies may provide novel functional targets to modulate RBC function, blood viscosity and flow, and tissue perfusion in disease conditions.

scholarly journals Homology modeling and functional characterization of multidrug effluxor Mta protein from Bacillus Atrophaeus: An explanatory insilico approach

2020 ◽  
Author(s):  
Mohammad Rejaur Rahman ◽  
Ishtiak Malique Chowdhury ◽  
Anik Banik ◽  
Emran Hossain Sajib
Keyword(s):  
Function Analysis ◽  
Functional Characterization ◽  
Bacillus Atrophaeus ◽  
Uncharacterized Protein ◽  
Protein Protein Interaction ◽  
Functional Aspects ◽  
Resistance Function ◽  
Spore Forming Bacteria ◽  
Protein Pocket

AbstractPhenotypically similar to B. subtilis, Bacillus atrophaeus is a Gram-positive, aerobic, spore-forming bacteria. It is a black-pigmented bacterial genus. Therefore, it is of interest to study the uncharacterized proteins in the genome. For a detailed computational sequence-structure-function analysis using available data and resources, an uncharacterized protein Mta (AKL87074.1) in the genome was selected. In this study, attempts were made to study the physicochemical properties, predict secondary structure, modeling the 3-D protein, pocket identification, protein-protein interaction and phylogenetic analysis of Mta protein. The predicted active site using CASTp is analyzed for understanding their multidrug resistance function. Because Mta is a MerR family member, these investigations on these functional aspects could lead us for better understanding of antibiotic resistance phenomenon.

Visfatin/PBEF/Nampt: structure, regulation and potential function of a novel adipokine

2008 ◽  
Vol 115 (1) ◽  
pp. 13-23 ◽  
Author(s):  
Grit Sommer ◽  
Antje Garten ◽  
Stefanie Petzold ◽  
Annette G. Beck-Sickinger ◽  
Matthias Blüher ◽  
...  
Keyword(s):  
Insulin Resistance ◽  
B Cell ◽  
Hormonal Regulation ◽  
Physiological Role ◽  
Nicotinamide Phosphoribosyltransferase ◽  
Insulin Mimetic ◽  
Enzymatic Function ◽  
Enhancing Factor ◽  
Structure Regulation ◽  
Cell Colony

Over the last few years, it has become obvious that obesity and insulin resistance are linked by a variety of proteins secreted by adipocytes. Visfatin/PBEF (pre-B-cell colony-enhancing factor) has recently been identified as a novel adipokine with insulin-mimetic effects. Furthermore, an enzymatic function has been reported that reveals visfatin/PBEF as Nampt (nicotinamide phosphoribosyltransferase; EC 2.4.2.12.). Moreover, reports on the structure and hormonal regulation of visfatin/PBEF/Nampt have given further insights into its potential physiological role. The present review summarizes studies on visfatin/PBEF/Nampt as a novel adipokine.

scholarly journals An NADH-Dependent Reductase from Eubacterium ramulus Catalyzes the Stereospecific Heteroring Cleavage of Flavanones and Flavanonols

2019 ◽  
Vol 85 (19) ◽  
Author(s):  
Annett Braune ◽  
Michael Gütschow ◽  
Michael Blaut
Keyword(s):  
Sequence Similarity ◽  
Bacterial Species ◽  
Functional Characterization ◽  
Intestinal Bacteria ◽  
Cyclic Ether ◽  
Content Type ◽  
Oxygen Sensitive ◽  
Key Enzyme ◽  
Catalyzed Reactions ◽  
Dietary Flavonoids

ABSTRACT The human intestinal anaerobe Eubacterium ramulus is known for its ability to degrade various dietary flavonoids. In the present study, we demonstrate the cleavage of the heterocyclic C-ring of flavanones and flavanonols by an oxygen-sensitive NADH-dependent reductase, previously described as enoate reductase, from E. ramulus. This flavanone- and flavanonol-cleaving reductase (Fcr) was purified following its heterologous expression in Escherichia coli and further characterized. Fcr cleaved the flavanones naringenin, eriodictyol, liquiritigenin, and homoeriodictyol. Moreover, the flavanonols taxifolin and dihydrokaempferol served as substrates. The catalyzed reactions were stereospecific for the (2R)-enantiomers of the flavanone substrates and for the (2S,3S)-configured flavanonols. The enantioenrichment of the nonconverted stereoisomers allowed for the determination of hitherto unknown flavanone racemization rates. Fcr formed the corresponding dihydrochalcones and hydroxydihydrochalcones in the course of an unusual reductive cleavage of cyclic ether bonds. Fcr did not convert members of other flavonoid subclasses, including flavones, flavonols, and chalcones, the latter indicating that the reaction does not involve a chalcone intermediate. This view is strongly supported by the observed enantiospecificity of Fcr. Cinnamic acids, which are typical substrates of bacterial enoate reductases, were also not reduced by Fcr. Based on the presence of binding motifs for dinucleotide cofactors and a 4Fe-4S cluster in the amino acid sequence of Fcr, a cofactor-mediated hydride transfer from NADH onto C-2 of the respective substrate is proposed. IMPORTANCE Gut bacteria play a crucial role in the metabolism of dietary flavonoids, thereby contributing to their activation or inactivation after ingestion by the human host. Thus, bacterial activities in the intestine may influence the beneficial health effects of these polyphenolic plant compounds. While an increasing number of flavonoid-converting gut bacterial species have been identified, knowledge of the responsible enzymes is still limited. Here, we characterized Fcr as a key enzyme involved in the conversion of flavonoids of several subclasses by Eubacterium ramulus, a prevalent human gut bacterium. Sequence similarity of this enzyme to hypothetical proteins from other flavonoid-degrading intestinal bacteria in databases suggests a more widespread occurrence of this enzyme. Functional characterization of gene products of human intestinal microbiota enables the assignment of metagenomic sequences to specific bacteria and, more importantly, to certain activities, which is a prerequisite for targeted modulation of gut microbial functionality.

scholarly journals Functional Characterization of Cholinergic Receptors in Melanoma Cells

Cancers ◽  
2020 ◽  
Vol 12 (11) ◽  
pp. 3141
Author(s):  
Anna Maria Lucianò ◽  
Ada Maria Tata
Keyword(s):  
Nervous System ◽  
Acetylcholine Receptors ◽  
Epithelial Mesenchymal Transition ◽  
Functional Characterization ◽  
Physiological Role ◽  
Melanoma Cells ◽  
Cholinergic Receptors ◽  
Mesenchymal Transition ◽  
Relevant Role

In the last two decades, the scientific community has come to terms with the importance of non-neural acetylcholine in light of its multiple biological and pathological functions within and outside the nervous system. Apart from its well-known physiological role both in the central and peripheral nervous systems, in the autonomic nervous system, and in the neuromuscular junction, the expression of the acetylcholine receptors has been detected in different peripheral organs. This evidence has contributed to highlight new roles for acetylcholine in various biological processes, (e.g., cell viability, proliferation, differentiation, migration, secretion). In addition, growing evidence in recent years has also demonstrated new roles for acetylcholine and its receptors in cancer, where they are involved in the modulation of cell proliferation, apoptosis, angiogenesis, and epithelial mesenchymal transition. In this review, we describe the functional characterization of acetylcholine receptors in different tumor types, placing attention on melanoma. The latest set of data accessible through literature, albeit limited, highlights how cholinergic receptors both of muscarinic and nicotinic type can play a relevant role in the migratory processes of melanoma cells, suggesting their possible involvement in invasion and metastasis.

scholarly journals Genomic and Transcriptome Analyses of a Thermophilic Bacterium Geobacillus stearothermophilus B5 Isolated from Compost Reveal Its Enzymatic Basis for Lignocellulose Degradation

2020 ◽  
Vol 8 (9) ◽  
pp. 1357
Author(s):  
Mengmeng Wang ◽  
Jiaxi Miao ◽  
Xuanqing Wang ◽  
Tuo Li ◽  
Han Zhu ◽  
...  
Keyword(s):  
Thermophilic Bacterium ◽  
Glycoside Hydrolases ◽  
Lignocellulose Degradation ◽  
Geobacillus Stearothermophilus ◽  
Carbohydrate Active Enzymes ◽  
Transcriptional Responses ◽  
Protein Coding ◽  
Clusters Of Orthologous Groups ◽  
Different Temperatures ◽  
Shock Proteins

A lignocellulose-degrading strain isolated from thermophilic compost was identified as Geobacillus stearothermophilus B5, and found able to secrete considerable amounts of enzymes at optimal temperature (60 °C) and pH (7.5). One circular contig of 3.37 Mbp was assembled from raw data, and 3371 protein-coding genes were predicted. Clusters of orthologous groups (COG) analysis revealed various genes with functions in polymeric substrate degradation, especially for Carbohydrate Active enZymes (CAZymes), such as glycoside hydrolases (GHs) and glycosyl transferases (GTs). Furthermore, the transcriptional responses of B5 at different temperatures—with rice straw provided as the sole carbon source—were analyzed. The results revealed that B5 could resist high temperature by upregulating heat shock proteins (HSPs), enhancing protein synthesis, and decreasing carbon catabolism. Briefly, B5 possesses the ability of lignocellulose degradation, and might be considered a potential inoculant for improving composting efficiency.

Identification of a bacterial di-haem cytochrome c peroxidase from Methylomicrobium album BG8

Microbiology ◽  
2010 ◽  
Vol 156 (9) ◽  
pp. 2682-2690 ◽  
Author(s):  
O. A. Karlsen ◽  
Ø. Larsen ◽  
H. B. Jensen
Keyword(s):  
Copper Ions ◽  
Sequence Similarity ◽  
Physiological Role ◽  
Growth Conditions ◽  
Inverse Pcr ◽  
Restriction Enzyme Digestion ◽  
Methylococcus Capsulatus ◽  
Gene Encoding ◽  
Reading Frame ◽  
Methylomicrobium Album

The nucleotide sequence of an open reading frame (corB) downstream of the copper-repressible CorA-encoding gene of the methanotrophic bacterium Methylomicrobium album BG8 was obtained by restriction enzyme digestion and inverse PCR. The amino acid sequence deduced from this gene showed significant sequence similarity to the surface-associated di-haem cytochrome c peroxidase (SACCP) previously isolated from Methylococcus capsulatus (Bath), including both c-type haem-binding motifs. Homology analysis placed this protein, phylogenetically, within the subfamily containing the M. capsulatus SACCP of the bacterial di-haem cytochrome c peroxidase (BCCP) family of proteins. Immunospecific recognition confirmed synthesis of the M. album CorB as a protein non-covalently associated with the outer membrane and exposed to the periplasm. corB expression is regulated by the availability of copper ions during growth and the protein is most abundant in M. album when grown at a low copper-to-biomass ratio, indicating an important physiological role of CorB under these growth conditions. corB was co-transcribed with the gene encoding CorA, constituting a copper-responding operon, which appears to be under the control of a σ 54-dependent promoter. M. album CorB is the second isolated member of the recently described subfamily of the BCCP family of proteins. So far, these proteins have only been described in methanotrophic bacteria.

Molecular cloning, chromosomal organization, and functional characterization of a sodium-dicarboxylate cotransporter from mouse kidney

2000 ◽  
Vol 279 (3) ◽  
pp. F482-F490 ◽  
Author(s):  
Ana M. Pajor ◽  
Nina N. Sun
Keyword(s):  
Sequence Similarity ◽  
Genomic Structure ◽  
Functional Characterization ◽  
Krebs Cycle ◽  
Initial Step ◽  
Xenopus Laevis Oocytes ◽  
Chromosome 11 ◽  
Inward Currents ◽  
Urinary Citrate

The sodium-dicarboxylate cotransporter of the renal proximal tubule, NaDC-1, reabsorbs filtered Krebs cycle intermediates and plays an important role in the regulation of urinary citrate concentrations.1 Low urinary citrate is a risk factor for the development of kidney stones. As an initial step in the characterization of NaDC-1 regulation, the genomic structure and functional properties of the mouse Na+-dicarboxylate cotransporter (mNaDC-1) were determined. The gene coding for mNaDC-1, Slc13a2, is found on chromosome 11. The gene is ∼24.9 kb in length and contains 12 exons. The mRNA coding for mNaDC-1 is found in kidney and small intestine. Expression of mNaDC-1 in Xenopus laevis oocytes results in increased transport of di- and tricarboxylates. The Michaelis-Menten constant ( K m) for succinate was 0.35 mM, and the K m for citrate was 0.6 mM. The transport of citrate was stimulated by acidic pH, whereas the transport of succinate was insensitive to pH changes. Transport by mNaDC-1 is electrogenic, and substrates produced inward currents in the presence of sodium. The sodium affinity was relatively high in mNaDC-1, with half-saturation constants for sodium of 10 mM (radiotracer experiments) and 28 mM at −50 mV (2-electrode voltage clamp experiments). Lithium acts as a potent inhibitor of transport, but it can also partially substitute for sodium. In conclusion, the mNaDC-1 is related in sequence and function to the other NaDC-1 orthologs. However, its function more closely resembles the rabbit and human orthologs rather than the rat NaDC-1, with which it shares higher sequence similarity.

Sign in / Sign up

Export Citation Format

Share Document