Cellulose is the most abundant polymer on Earth and the main component of plant biomass. It normally presents itself in fibres associated to other biopolymers, namely, hemicelluloses and lignin, in a complex structural matrix in plant cell walls [1]. This complex matrix limits the extent and rate of utilization of plant biomass, usually requiring harsh pre-treatments and the action of multiple enzymes to perform the full breakdown of the structure [2]. Although multiple enzymatic activities are responsible of the conversion of cellulose into simpler molecules, collectively known as cellulases, β-1-4-endoglucanases (EC 3.2.1.4) are especially important as they act on the cellulose chain cleaving internal glycosidic bonds and releasing oligosaccharides of different lengths. These are further hydrolysed by other cellulolytic enzymes, such as non-reducing end cellobiohydrolases (EC 3.2.1.91), reducing-end cellobiohydrolases (EC 3.2.1.176) and cellodextrinases (EC 3.2.1.74), and finally converted in glucose by β-glucosidases (EC 3.2.1.21) reducing the product inhibition of all the enzymes mentioned before [3]. Endoglucanases have been classified along with other enzymes based on sequence similarity in the Carbohydrate Active Enzymes (CAZy) Database [4] ( ) in 12 Glycosyl Hydrolase families: GH5, GH6, GH7, GH8, GH9, GH12, GH44, GH45, GH48, GH51, GH74, and GH124. Endoglucanases have multiple biotechnological applications, and are especially important in the valorisation of agro-industrial by-products [2] and in the production of biofuels combined with β-glucosidases to produce glucose later fermented into (bio)ethanol [5], as these two applications are directly targeting environmental challenges. In the textile industry there are several enzymatic processes, such as biostoning (to give a wash-down look on cotton clothes) and biopolishing (softening and brightening of cotton surfaces) that remove cellulose fibres and replace more harsh treatments [6]. Similarly, detergent formulations can include endoglucanases, also brightening and softening cotton fabrics [7]. They are also employed in the food and brewing industries, improving digestibility of food and decreasing viscosity, and increasing fermentable compounds for the elaboration of alcoholic drinks [6]. These properties have also been exploited in the animal feed industry, enhancing digestibility and nutrient bioavailability [6]. The pulp and paper industry has many uses for endoglucanases including biopulping, treatment of pulp wastes, deinking and removal of pollutants from paper [6]. Other reported uses include waste management, improvement of soils for agriculture and extraction of bioactive compounds, pigments and oils from plants [3]. Many of these applications are benefited from the use of combinations of various enzymes (enzyme cocktails) or multifunctional enzymes, as the plant biomass is composed of a complex matrix of cellulose, hemicelluloses and lignin [8]. Industrial processes such as biofuel production, food processing, treatments in the pulp and paper industry and production of nutraceuticals have been explored in this context, among others [9, 10]. In this biotechnological context, cellulases from thermophilic microorganisms have added advantages over their mesophilic counterparts. First of all, they are able to withstand the harsh conditions associated with industrial processes. This in turn reduces costs related to the need of cooling large amounts of water or other solvents. Moreover, the diffusion rates and solubility of reagents are higher at high temperatures and the risks of contamination are reduced [11]. Sources for these thermophilic enzymes are varied and included habitats, such as terrestrial hot springs, hydrothermal vents, compost and hydrocarbon reservoirs, among others [12]. Of these, terrestrial hot springs are one of the most common sources of thermophilic enzymes ([13], since the pioneering work describing the first thermophilic organisms in Yellowstone National Park (USA) [14] and the isolation of the Thermus aquaticus polymerase that allowed the development of the PCR technique. As with many extremophilic microorganisms, thermophiles are difficult to grow in laboratory conditions and culture independent methods such as metagenomics are needed for assessing their metabolic potential [15]. Lists of thermophilic cellulases found by metagenomics [12] and characterized thermophilic cellulases [3] are available, and novel cellulases found following this strategy continue to be discovered [16, 17] showing the interest for these biocatalysts. Moreover, the discovery of multifunctional enzymes that can act on more than one biomass polymer [18,19,20], and the characterization of microbial consortiums that produce multiple lignocellulolytic enzymes [21] are also recent research focuses. The development of high-throughput Next Generation Sequencing (NGS) technologies, and more specifically of shotgun metagenomics, has allowed to directly sequence the large number of genomes present in environmental samples, using multiple templates in parallel without targeting specific genes [15, 22]. The method relies on annotated data (reference genomes and gene and protein databases), as gene search and functional annotation using bioinformatic tools is based on alignment and homology to deposited sequences [15, 22, 23, 24]. Nevertheless, sequence based metagenomic studies also face various challenges, including quality and length of the reads generated, and amplification bias and other artifacts, such as chimeric sequences and secondary structures [22]. It is also important to consider that the success of the method heavily relies on the quality of the database annotation and is limited to find somewhat similar sequences in known protein families [23]. When the objective is to bioprospect for novel gene products, the short reads can be linked together into a bigger sequence (contig). Due to the high computational demand of methods based on overlapping reads, many de novo assemblers use instead a de Bruijn graph approach [24]. Predicted Open Reading Frames (ORF) within the reconstructed contigs can then be submitted and aligned to known sequences deposited in annotated databases, allowing the identification of ecological or biotechnological functions of interest. Finally, is important to remark that functional characterization of predicted gene products is still necessary to confirm the results of the in silico analysis [12]. In this regard, the selection of the heterologous expression system has become increasingly important in the context of biotechnological driven bioprospections, as factors such as thermostability, purification from intracellular or extracellular medium and enzyme kinetics are all affected by it [25]. Differences such as the ability to perform post-translational modifications including glycosylation, and high levels of enzymatic yield make hosts such as the yeast S. cerevisiae attractive for the expression of recombinant thermophilic enzymes [1,2,3,4,5, 7,8,9,10, 12, 16,17,18,19,20,2126].
The most common adverse events attributed to DFX therapy are gastrointestinal disturbances and skin rash (Table 3). Diarrhea is more common in the elderly. Mild, nonprogressive increases in serum creatinine and liver enzyme levels have also be noted. Recommendations for their monitoring and management are summarized in Table 3. A boxed warning was added to the United States DFX prescribing information, although this amendment has not been adopted by the European Health Authority or applied globally. The warning indicates that DFX may cause renal and hepatic impairment, including failure, and gastrointestinal hemorrhage. In some reported cases, these reactions were fatal. However, these reactions were observed in patients with advanced age, high-risk MDS, and those with underlying renal or hepatic impairment or low platelet counts. DFX is contraindicated in patients with renal and hepatic failure.81
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Figure 1 shows the search strategy identified 6981 articles from the listed databases; articles were downloaded to EndNote to remove a total of 3590 duplicates. Upon screening for inclusion criteria in titles and abstracts, the full text of 86 articles was independently reviewed. After review using the exclusion criteria, 19 articles were included for analysis. The additional record search resulted in two additional articles for a final total of 21. 2ff7e9595c
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