Structural and functional properties of the hemoglobin components from the Caspian Sea Acipenser persicus and Acipenser stellatus

Hemoglobin (Hb) variability is a commonly used index of phylogenetic differentiation and molecular adaptation in fish. In the current study, the structural and functional characteristics of Hbs from two Sturgeon species of the Southern Caspian Sea Basin were investigated. After extraction and separation of hemoglobin from whole blood , the polyacrylamide gel electrophoresis (SDSPAGE), native-PAGE and isoelectric focusing (IEF) were used to confirm Hb variability in these fishes. Ion-exchange on CM-cellulose chromatography was used for purification of the dominant Hbs from these fishes. The accuracy of the methods was confirmed by IEF and SDS-PAGE. Spectral studies using fluorescence spectrophotometery, circular dichroism spectropolarimetry (CD) analysis and UV–vis spectrophotometery. Oxygen affinities of these Hbs were compared using Hb-oxygen dissociation curves. Also, the dominant Hbs from these blood fishes were utilized for further experiments. The behavior of Hbs during the denaturation process by n-dodecyl trimethylammonium bromide (DTAB) is investigated by UV–vis spectrophotometer and circular dichroism spectropolarimetry. The thermal denaturation properties of the Hbs wereinvestigated by differential scanning calorimetry (DSC) and Hbs aggregation performed chemically in the presence of dithiotreitol (DTT) by UV–vis spectrophotometer and chemometric study. The results demonstrate a significant relationship between stability of fish hemoglobins and the ability of fish for entering to deeper depths. The UV–Vis absorption spectra identified species of hemoglobin and showed the concentration of oxyHb and metHb decreases and deoxyHb increases upon interaction with DTAB. Besides the UV–vis spectrophotometry, the interaction of DTAB with hemoglobins has been studied using circular dichroism spectropolarimetry analysis. This experiment was utilized to measure the unfolding mechanism and compared alpha-helix secondary structure under different conditions for Hbs. The results reveal that the Acipenser stellatus Hb in comparison with Acipenser persicus Hb has more stability and more structural compactness. Besides, the results confirm the hypothesis that there is a meaningful relation between average habitat depth, partial oxygen pressure, oxygen affinity, structural compactness of Hb, and its stability

A metagenomic catalog for exploring the plastizymes landscape covering taxa, genes, and proteins

Abstract There are significant environmental and health concerns associated with the current inefficient plastic recycling process. This study presents the first integrated reference catalog of plastic-contaminated environments obtained using an insilico workflow that could play a significant role in discovering new plastizymes. Here, we combined 66 whole metagenomic data from plastic-contaminated environment samples from four previously collected metagenome data with our new sample. In this study, an integrated plastic-contaminated environment gene, protein, taxa, and plastic degrading enzyme catalog (PDEC) was constructed. These catalogs contain 53,300,583 non-redundant genes and proteins, 691 metagenome-assembled genomes, and 136,654 plastizymes. Based on KEGG and eggNOG annotations, 42% of recognized genes lack annotations, indicating their functions remain elusive and warrant further investigation. Additionally, the PDEC catalog highlights hydrolases, peroxidases, and cutinases as the prevailing plastizymes. Ultimately, following multiple validation procedures, our effort focused on pinpointing enzymes that exhibited the highest similarity to the introduced plastizymes in terms of both sequence and three-dimensional structural aspects. This encompassed evaluating the linear composition of constituent units as well as the complex spatial conformation of the molecule. The resulting catalog is expected to improve the resolution of future multi-omics studies, providing new insights into plastic-pollution related research

Efficient bioconversion of lignocellulosic waste by a novel computationally screened hyperthermostable enzyme from a specialized microbiota

A large amount of lignocellulosic waste is generated every day in the world, and their accumulation in the agroecosystems, integration in soil compositions, or incineration for energy production has severe environmental pollution effects. Using enzymes as biocatalysts for the biodegradation of lignocellulosic materials, especially in harsh processing conditions, is a practical step towards green energy and environmental biosafety. Hence, the current study focuses on enzyme computationally screened from camel rumen metagenomics data as specialized microbiota that have the capacity to degrade lignocellulosic-rich and recalcitrant materials. The novel hyperthermostable xylanase named PersiXyn10 with the performance at extreme conditions was proper activity within a broad temperature (30-100 ℃) and pH range (4.0-11.0) but showed the maximum xylanolytic activity in severe alkaline and temperature conditions, pH 8.0 and temperature 90 ℃. Also, the enzyme had highly resistant to metals, surfactants, and organic solvents in optimal conditions. The introduced xylanase had unique properties in terms of thermal stability by maintaining over 82% of its activity after 15 days of incubation at 90 ℃. Considering the crucial role of hyperthermostable xylanases in the paper industry, the PersiXyn10 was subjected to biodegradation of paper pulp. The proper performance of hyperthermostable PersiXyn10 on the paper pulp was confirmed by structural analysis (SEM and FTIR) and produced 31.64 g/L of reducing sugar after 144 h hydrolysis. These results proved the applicability of the hyperthermostable xylanase in biobleaching and saccharification of lignocellulosic biomass for declining the environmental hazards

A computational method for prediction of xylanase enzymes activity in strains of Bacillus subtilis based on pseudo amino acid composition features.

Xylanases are hydrolytic enzymes which based on physicochemical properties, structure, mode of action and substrate specificities are classified into various glycoside hydrolase (GH) families. The purpose of this study is to show that the activity of the members of the xylanase family in the specified pH and temperature conditions can be computationally predicted. The proposed computational regression model was trained and tested with the Pseudo Amino Acid Composition (PseAAC) features extracted solely from the amino acid sequences of enzymes. The xylanases with experimentally determined activities were used as the training dataset to adjust the model parameters. To develop the model, 41 strains of Bacillus subtilis isolated from field soil were screened. From them, 28 strains with the highest halo diameter were selected for further studies. The performance of the model for prediction of xylanase activity was evaluated in three different temperature and pH conditions using stratified cross-validation and jackknife methods. The trained model can be used for determining the activity of newly found xylanases in the specified condition. Such computational models help to scale down the experimental costs and save time by identifying enzymes with appropriate activity for scientific and industrial usage. Our methodology for activity prediction of xylanase enzymes can be potentially applied to the members of the other enzyme families. The availability of sufficient experimental data in specified pH and temperature conditions is a prerequisite for training the learning model and to achieve high accuracy

Enhancing the ethanol production by exploiting a novel metagenomic-derived bifunctional xylanase/β-glucosidase enzyme with improved β-glucosidase activity by a nanocellulose carrier

Some enzymes can catalyze more than one chemical conversion for which they are physiologically specialized. This secondary function, which is called underground, promiscuous, metabolism, or cross activity, is recognized as a valuable feature and has received much attention for developing new catalytic functions in industrial applications. In this study, a novel bifunctional xylanase/β-glucosidase metagenomic-derived enzyme, PersiBGLXyn1, with underground β-glucosidase activity was mined by in-silico screening. Then, the corresponding gene was cloned, expressed and purified. The PersiBGLXyn1 improved the degradation efficiency of organic solvent pretreated coffee residue waste (CRW), and subsequently the production of bioethanol during a separate enzymatic hydrolysis and fermentation (SHF) process. After characterization, the enzyme was immobilized on a nanocellulose (NC) carrier generated from sugar beet pulp (SBP), which remarkably improved the underground activity of the enzyme up to four-fold at 80°C and up to two-fold at pH 4.0 compared to the free one. The immobilized PersiBGLXyn1 demonstrated 12 to 13-fold rise in half-life at 70 and 80°C for its underground activity. The amount of reducing sugar produced from enzymatic saccharification of the CRW was also enhanced from 12.97 g/l to 19.69 g/l by immobilization of the enzyme. Bioethanol production was 29.31 g/l for free enzyme after 72 h fermentation, while the immobilized PersiBGLXyn1 showed 51.47 g/l production titre. Overall, this study presented a cost-effective in-silico metagenomic approach to identify novel bifunctional xylanase/β-glucosidase enzyme with underground β-glucosidase activity. It also demonstrated the improved efficacy of the underground activities of the bifunctional enzyme as a promising alternative for fermentable sugars production and subsequent value-added products

Highly Efficient Computationally Derived Novel Metagenome α-Amylase With Robust Stability Under Extreme Denaturing Conditions

α-Amylases are among the very critical enzymes used for different industrial purposes. Most α-amylases cannot accomplish the requirement of industrial conditions and easily lose their activity in harsh environments. In this study, a novel α-amylase named PersiAmy1 has been identified through the multistage in silico screening pipeline from the rumen metagenomic data. The long-term storage of PersiAmy1 in low and high temperatures demonstrated 82.13 and 71.01% activities after 36 days of incubation at 4 and 50°C, respectively. The stable α-amylase retained 61.09% of its activity after 180 min of incubation at 90°C and was highly stable in a broad pH range, showing 60.48 and 86.05% activities at pH 4.0 and pH 9.0 after 180 min of incubation, respectively. Also, the enzyme could resist the high-salinity condition and demonstrated 88.81% activity in the presence of 5 M NaCl. PersiAmy1 showed more than 74% activity in the presence of various metal ions. The addition of the detergents, surfactants, and organic solvents did not affect the α-amylase activity considerably. Substrate spectrum analysis showed that PersiAmy1 could act on a wide array of substrates. PersiAmy1 showed high stability in inhibitors and superb activity in downstream conditions, thus useful in detergent and baking industries. Investigating the applicability in detergent formulation, PersiAmy1 showed more than 69% activity after incubation with commercial detergents at different temperatures (30–50°C) and retained more than 56% activity after incubation with commercial detergents for 3 h at 10°C. Furthermore, the results of the wash performance analysis exhibited a good stain removal at 10°C. The power of PersiAmy1 in the bread industry revealed soft, chewable crumbs with improved volume and porosity compared with control. This study highlights the intense power of robust novel PersiAmy1 as a functional bio-additive in many industrial applications

Image_1_Enhancing the ethanol production by exploiting a novel metagenomic-derived bifunctional xylanase/β-glucosidase enzyme with improved β-glucosidase activity by a nanocellulose carrier.JPEG

Some enzymes can catalyze more than one chemical conversion for which they are physiologically specialized. This secondary function, which is called underground, promiscuous, metabolism, or cross activity, is recognized as a valuable feature and has received much attention for developing new catalytic functions in industrial applications. In this study, a novel bifunctional xylanase/β-glucosidase metagenomic-derived enzyme, PersiBGLXyn1, with underground β-glucosidase activity was mined by in-silico screening. Then, the corresponding gene was cloned, expressed and purified. The PersiBGLXyn1 improved the degradation efficiency of organic solvent pretreated coffee residue waste (CRW), and subsequently the production of bioethanol during a separate enzymatic hydrolysis and fermentation (SHF) process. After characterization, the enzyme was immobilized on a nanocellulose (NC) carrier generated from sugar beet pulp (SBP), which remarkably improved the underground activity of the enzyme up to four-fold at 80°C and up to two-fold at pH 4.0 compared to the free one. The immobilized PersiBGLXyn1 demonstrated 12 to 13-fold rise in half-life at 70 and 80°C for its underground activity. The amount of reducing sugar produced from enzymatic saccharification of the CRW was also enhanced from 12.97 g/l to 19.69 g/l by immobilization of the enzyme. Bioethanol production was 29.31 g/l for free enzyme after 72 h fermentation, while the immobilized PersiBGLXyn1 showed 51.47 g/l production titre. Overall, this study presented a cost-effective in-silico metagenomic approach to identify novel bifunctional xylanase/β-glucosidase enzyme with underground β-glucosidase activity. It also demonstrated the improved efficacy of the underground activities of the bifunctional enzyme as a promising alternative for fermentable sugars production and subsequent value-added products.</p