Thermobifida fusca
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA35093 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| Other | ||||
| AOSG01.dat.gz | Other | |||
| AOSG01.fasta.gz | Fasta.gz | |||
| AOSG01.master.dat | Other | |||
| BBGT01.dat.gz | Other |
Items per page: 5 1 - 5 of 47 |
Similar Datasets
Project description:Fungus that produces thermostable enzymes
| PRJNA32777 | ENA
Project description:Thermophilic bacterium that produces thermostable enzymes of use in industry
| PRJNA10617 | ENA
Project description:Thermobifida fusca is of biotechnological interest due to its ability to produce an array of plant cell wall hydrolytic enzymes. Nonetheless, only one T. fusca bacteriophage with genome information has been reported to date. This study was aimed at discovering more relevant bacteriophages to expand the existing knowledge of phage diversity for this host species. With this end in view, a thermostable T. fusca bacteriophage P318, which belongs to the Siphoviridae family, was isolated and characterized. P318 has a double-stranded DNA genome of 48,045 base pairs with 3'-extended COS ends, on which 52 putative ORFs are organized into clusters responsible for the order of genome replication, virion morphogenesis, and the regulation of the lytic/lysogenic cycle. In comparison with T. fusca and the previously discovered bacteriophage P1312, P318 has a much lower G+C content in its genome except at the region encompassing ORF42, which produced a protein with unknown function. P1312 and P318 share very few similarities in their genomes except for the regions encompassing ORF42 of P318 and ORF51 of P1312 that are homologous. Thus, acquisition of ORF42 by lateral gene transfer might be an important step in the evolution of P318.
| S-EPMC6893835 | biostudies-literature
Project description:Actinomycetes have a long history of being the source of numerous valuable natural products and medicinals. To expedite product discovery and optimization of biochemical production, high-throughput technologies can now be used to screen the library of compounds present (or produced) at a given time in an organism. This not only facilitates chemical product screening, but also provides a comprehensive methodology to the study cellular metabolic networks to inform cellular engineering. Here, we present some of the first metabolomic data of the industrial cellulolytic actinomycete Thermobifida fusca generated using LC-MS/MS. The underlying objective of conducting global metabolite profiling was to gain better insight on the innate capabilities of T. fusca, with a long-term goal of facilitating T. fusca-based bioprocesses. The T. fusca metabolome was characterized for growth on two cellulose-relevant carbon sources, cellobiose and Avicel. Furthermore, the comprehensive list of measured metabolites was computationally integrated into a metabolic model of T. fusca, to study metabolic shifts in the network flux associated with carbohydrate and amino acid metabolism.
| S-EPMC5746737 | biostudies-literature
Project description:A relationship between processivity and synergism has not been reported for cellulases, although both characteristics are very important for hydrolysis of insoluble substrates. Mutation of two residues located in the active site tunnel of Thermobifida fusca exocellulase Cel6B increased processivity on filter paper. Surprisingly, mixtures of the Cel6B mutant enzymes and T. fusca endocellulase Cel5A did not show increased synergism or processivity, and the mutant enzyme which had the highest processivity gave the poorest synergism. This study suggests that improving exocellulase processivity might be not an effective strategy for producing improved cellulase mixtures for biomass conversion. The inverse relationship between the activities of many of the mutant enzymes with bacterial microcrystalline cellulose and their activities with carboxymethyl cellulose indicated that there are differences in the mechanisms of hydrolysis for these substrates, supporting the possibility of engineering Cel6B to target selected substrates.
| S-EPMC2772456 | biostudies-literature
Project description:Type I Baeyer-Villiger monooxygenases (BVMOs) strongly prefer NADPH over NADH as an electron donor. In order to elucidate the molecular basis for this coenzyme specificity, we have performed a site-directed mutagenesis study on phenylacetone monooxygenase (PAMO) from Thermobifida fusca. Using sequence alignments of type I BVMOs and crystal structures of PAMO and cyclohexanone monooxygenase in complex with NADP(+), we identified four residues that could interact with the 2'-phosphate moiety of NADPH in PAMO. The mutagenesis study revealed that the conserved R217 is essential for binding the adenine moiety of the nicotinamide coenzyme while it also contributes to the recognition of the 2'-phosphate moiety of NADPH. The substitution of T218 did not have a strong effect on the coenzyme specificity. The H220N and H220Q mutants exhibited a ~3-fold improvement in the catalytic efficiency with NADH while the catalytic efficiency with NADPH was hardly affected. Mutating K336 did not increase the activity of PAMO with NADH, but it had a significant and beneficial effect on the enantioselectivity of Baeyer-Villiger oxidations and sulfoxidations. In conclusion, our results indicate that the function of NADPH in catalysis cannot be easily replaced by NADH. This finding is in line with the complex catalytic mechanism and the vital role of the coenzyme in BVMOs.
| S-EPMC2956058 | biostudies-literature
Project description:The Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-CRISPR-associated (Cas) defense system is the only adaptive and inheritable immunity found in prokaryotes. The immunity is achieved through a multistep process of adaptation, expression, and interference. In the Type I-E system, interference is mediated by the CRISPR-associated complex for antiviral defense (Cascade), which recognizes invading double-stranded DNA (dsDNA) through the protospacer adjacent motif (PAM) by one of the Cascade components, Cse1. Here, we report the crystal structure of Thermobifida fusca Cse1 at 3.3 Å resolution. T. fusca Cse1 reveals the chair-like two-domain architecture with a well-defined flexible loop, L1, located at the larger N-terminal domain, which was not observed in previous structures of the single Cse1 protein. Structure-based mutagenesis analysis demonstrates that the well-defined flexible loop and a partially conserved structural motif ([FW]-X-[TH]) are involved in PAM binding and recognition, respectively. Moreover, structural docking of T. fusca Cse1 into Escherichia coli Cascade cryoelectron microscopy maps, coupled with structural comparison, reveals a conserved positive patch that is contiguous with Cse2 in the Cascade complex and adjacent to the Cas3 binding site, suggesting its role in R-loop formation/stabilization and the recruitment of Cas3 for target cleavage. Consistent with the structural observation, the introduction of alanine mutations at this positive patch abolished DNA binding activity by Cse1. Taken together, these results suggest that Cse1 is a critical Cascade component involved in Cascade assembly, dsDNA target recognition, R-loop formation, and Cas3 recruitment for target cleavage.
| S-EPMC4315661 | biostudies-literature
Project description:Thermostable enzymes employ various structural features dictated at the amino-acid sequence level that allow them to maintain their integrity at higher temperatures. Many hypotheses as to the nature of thermal stability have been proposed, including optimized core hydrophobicity and an increase in charged surface residues to enhance polar solvent interactions for solubility. Here, the three-dimensional structure of the family GH11 xylanase from the moderate thermophile Thermobifida fusca in its trapped covalent glycosyl-enzyme intermediate complex is presented. Interactions with the bound ligand show fewer direct hydrogen bonds from ligand to protein than observed in previous complexes from other species and imply that binding of the xylan substrate involves several water-mediated hydrogen bonds.
| S-EPMC3274389 | biostudies-literature
Project description:Thermobifida fusca Cel9A-90 is a processive endoglucanase consisting of a family 9 catalytic domain (CD), a family 3c cellulose binding module (CBM3c), a fibronectin III-like domain, and a family 2 CBM. This enzyme has the highest activity of any individual T. fusca enzyme on crystalline substrates, particularly bacterial cellulose (BC). Mutations were introduced into the CD or the CBM3c of Cel9A-68 using site-directed mutagenesis. The mutant enzymes were expressed in Escherichia coli; purified; and tested for activity on four substrates, ligand binding, and processivity. The results show that H125 and Y206 play an important role in activity by forming a hydrogen bonding network with the catalytic base, D58; another important supporting residue, D55; and Glc(-1) O1. R378, a residue interacting with Glc(+1), plays an important role in processivity. Several enzymes with mutations in the subsites Glc(-2) to Glc(-4) had less than 15% activity on BC and markedly reduced processivity. Mutant enzymes with severalfold-higher activity on carboxymethyl cellulose (CMC) were found in the subsites from Glc(-2) to Glc(-4). The CBM3c mutant enzymes, Y520A, R557A/E559A, and R563A, had decreased activity on BC but had wild-type or improved processivity. Mutation of D513, a conserved residue at the end of the CBM, increased activity on crystalline cellulose. Previous work showed that deletion of the CBM3c abolished crystalline activity and processivity. This study shows that it is residues in the catalytic cleft that control processivity while the CBM3c is important for loose binding of the enzyme to the crystalline cellulose substrate.
| S-EPMC1907127 | biostudies-literature
Project description:F420H2-dependent enzymes reduce a wide range of substrates that are otherwise recalcitrant to enzyme-catalyzed reduction, and their potential for applications in biocatalysis has attracted increasing attention. Thermobifida fusca is a moderately thermophilic bacterium and holds high biocatalytic potential as a source for several highly thermostable enzymes. We report here on the isolation and characterization of a thermostable F420: NADPH oxidoreductase (Tfu-FNO) from T. fusca, the first F420-dependent enzyme described from this bacterium. Tfu-FNO was heterologously expressed in Escherichia coli, yielding up to 200 mg of recombinant enzyme per liter of culture. We found that Tfu-FNO is highly thermostable, reaching its highest activity at 65 °C and that Tfu-FNO is likely to act in vivo as an F420 reductase at the expense of NADPH, similar to its counterpart in Streptomyces griseus We obtained the crystal structure of FNO in complex with NADP+ at 1.8 Å resolution, providing the first bacterial FNO structure. The overall architecture and NADP+-binding site of Tfu-FNO were highly similar to those of the Archaeoglobus fulgidus FNO (Af-FNO). The active site is located in a hydrophobic pocket between an N-terminal dinucleotide binding domain and a smaller C-terminal domain. Residues interacting with the 2'-phosphate of NADP+ were probed by targeted mutagenesis, indicating that Thr-28, Ser-50, Arg-51, and Arg-55 are important for discriminating between NADP+ and NAD+ Interestingly, a T28A mutant increased the kinetic efficiency >3-fold as compared with the wild-type enzyme when NADH is the substrate. The biochemical and structural data presented here provide crucial insights into the molecular recognition of the two cofactors, F420 and NAD(P)H by FNO.
| S-EPMC5473218 | biostudies-literature