An amylopullulanase (ApuNP1) from Geobacillus thermoleovorans NP1: biochemical characterization and its potential industrial applications

2019 ◽  
Vol 49 (2) ◽  
pp. 127-135
Author(s):  
Nihan Arabacı ◽  
Burhan Arıkan
Keyword(s):  
Biochemical Characterization ◽  
Industrial Applications ◽  
Geobacillus Thermoleovorans

scholarly journals Structure-based enzyme engineering improves donor-substrate recognition of Arabidopsis thaliana glycosyltransferases

2020 ◽  
Vol 477 (15) ◽  
pp. 2791-2805
Author(s):  
Aishat Akere ◽  
Serena H. Chen ◽  
Xiaohan Liu ◽  
Yanger Chen ◽  
Sarath Chandra Dantu ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Biochemical Characterization ◽  
Structural Information ◽  
Substrate Recognition ◽  
Industrial Applications ◽  
Enzyme Engineering ◽  
Donor Substrate ◽  
Substrate Preferences ◽  
Dynamics Simulations ◽  
Experimental Crystal

Glycosylation of secondary metabolites involves plant UDP-dependent glycosyltransferases (UGTs). UGTs have shown promise as catalysts in the synthesis of glycosides for medical treatment. However, limited understanding at the molecular level due to insufficient biochemical and structural information has hindered potential applications of most of these UGTs. In the absence of experimental crystal structures, we employed advanced molecular modeling and simulations in conjunction with biochemical characterization to design a workflow to study five Group H Arabidopsis thaliana (76E1, 76E2, 76E4, 76E5, 76D1) UGTs. Based on our rational structural manipulation and analysis, we identified key amino acids (P129 in 76D1; D374 in 76E2; K275 in 76E4), which when mutated improved donor substrate recognition than wildtype UGTs. Molecular dynamics simulations and deep learning analysis identified structural differences, which drive substrate preferences. The design of these UGTs with broader substrate specificity may play important role in biotechnological and industrial applications. These findings can also serve as basis to study other plant UGTs and thereby advancing UGT enzyme engineering.

scholarly journals Biochemical Characterization of Cellulase From Bacillus subtilis Strain and its Effect on Digestibility and Structural Modifications of Lignocellulose Rich Biomass

2021 ◽  
Vol 9 ◽  
Author(s):  
Waseem Ayoub Malik ◽  
Saleem Javed
Keyword(s):  
Bacillus Subtilis ◽  
Biochemical Characterization ◽  
Biomass Conversion ◽  
Industrial Applications ◽  
Partial Purification ◽  
Subtilis Strain ◽  
Complex Nature ◽  
Structural Modifications ◽  
Optimum Activity

Microbial cellulases have become the mainstream biocatalysts due to their complex nature and widespread industrial applications. The present study reports the partial purification and characterization of cellulase from Bacillus subtilis CD001 and its application in biomass saccharification. Out of four different substrates, carboxymethyl cellulose, when amended as fermentation substrate, induced the highest cellulase production from B. subtilis CD001. The optimum activity of CMCase, FPase, and amylase was 2.4 U/ml, 1.5 U/ml, and 1.45 U/ml, respectively. The enzyme was partially purified by (NH4)2SO4 precipitation and sequenced through LC-MS/MS. The cellulase was found to be approximately 55 kDa by SDS-PAGE and capable of hydrolyzing cellulose, as confirmed by zymogram analysis. The enzyme was assigned an accession number AOR98335.1 and displayed 46% sequence homology with 14 peptide-spectrum matches having 12 unique peptide sequences. Characterization of the enzyme revealed it to be an acidothermophilic cellulase, having an optimum activity at pH 5 and a temperature of 60°C. Kinetic analysis of partially purified enzyme showed the Km and Vmax values of 0.996 mM and 1.647 U/ml, respectively. The enzyme activity was accelerated by ZnSO4, MnSO4, and MgSO4, whereas inhibited significantly by EDTA and moderately by β-mercaptoethanol and urea. Further, characterization of the enzyme saccharified sugarcane bagasse, wheat straw, and filter paper by SEM, ATR-FTIR, and XRD revealed efficient hydrolysis and structural modifications of cellulosic materials, indicating the potential industrial application of the B. subtilis CD001 cellulase. The findings demonstrated the potential suitability of cellulase from B. subtilis CD001 for use in current mainstream biomass conversion into fuels and other industrial processes.

scholarly journals Biochemical Characterization of the Amylase Activity from the New Haloarchaeal Strain Haloarcula sp. HS Isolated in the Odiel Marshlands

Biology ◽  
2021 ◽  
Vol 10 (4) ◽  
pp. 337
Author(s):  
Patricia Gómez-Villegas ◽  
Javier Vigara ◽  
Luis Romero ◽  
Cecilia Gotor ◽  
Sara Raposo ◽  
...  
Keyword(s):  
Biochemical Characterization ◽  
Amylase Activity ◽  
Extreme Environments ◽  
Three Dimensional ◽  
Large Family ◽  
Industrial Applications ◽  
Dimensional Structure ◽  
Glycosyl Hydrolases ◽  
Calcium Dependent ◽  
Alpha Amylases

Alpha-amylases are a large family of α,1-4-endo-glycosyl hydrolases distributed in all kingdoms of life. The need for poly-extremotolerant amylases encouraged their search in extreme environments, where archaea become ideal candidates to provide new enzymes that are able to work in the harsh conditions demanded in many industrial applications. In this study, a collection of haloarchaea isolated from Odiel saltern ponds in the southwest of Spain was screened for their amylase activity. The strain that exhibited the highest activity was selected and identified as Haloarcula sp. HS. We demonstrated the existence in both, cellular and extracellular extracts of the new strain, of functional α-amylase activities, which showed to be moderately thermotolerant (optimum around 60 °C), extremely halotolerant (optimum over 25% NaCl), and calcium-dependent. The tryptic digestion followed by HPLC-MS/MS analysis of the partially purified cellular and extracellular extracts allowed to identify the sequence of three alpha-amylases, which despite sharing a low sequence identity, exhibited high three-dimensional structure homology, conserving the typical domains and most of the key consensus residues of α-amylases. Moreover, we proved the potential of the extracellular α-amylase from Haloarcula sp. HS to treat bakery wastes under high salinity conditions.

scholarly journals Degradation Patterns of GH11 xylanases for Efficiently Hydrolyzing xylan in Aspergillus Niger An76

2021 ◽  
Author(s):  
Shu Zhang ◽  
Sha Zhao ◽  
Weihao Shang ◽  
Xiuyun Wu ◽  
Yingjie Li ◽  
...  
Keyword(s):  
Aspergillus Niger ◽  
Filamentous Fungi ◽  
Biochemical Characterization ◽  
Catalytic Performance ◽  
Industrial Applications ◽  
Transcriptional Analysis ◽  
Degrading Enzymes ◽  
Beechwood Xylan ◽  
Catalytic Functions ◽  
Encoding Genes

Abstract Background: Xylan is the most abundant hemicellulose polysaccharide in nature. Endo-xylanases from GH10 and GH11 families are the most critical xylan degrading enzymes. Filamentous fungi are highly effective xylan degraders and possess numerous xylan degrading isoenzyme-encoding genes, especially Aspergillus niger. Most noteworthy, the amplification of the GH11 xylanase-encoding genes occurs frequently in an organism, but the knowledge of each GH11 xylanases is little known. Results: A. niger An76 encoded a comprehensive set of xylan-degrading enzymes, including five endo-xylanases (one GH10 and four GH11). Quantitative transcriptional analysis showed that three xylanases were up-regulated by xylose substrates, and the order and amount of enzyme secretion differed. Specifically, XynA and XynB were initially secreted successively, followed by XynC. Structural bioinformatics analysis indicated that the different modes of action of the three GH11 xylanases may be due to intricate hydrogen bonding between substrates and functional residues in the active site architectures. Heterologous expression and biochemical characterization of three GH11 xylanases (XynA, XynB and XynD) revealed differences in catalytic performance and product profiles. Furthermore, XynA and XynB displayed obvious synergistic action against beechwood xylan. Conclusions: We investigated subtle differences in the functions of different isoenzymes in the same family using a combination of physiological and biochemical experiments. The transcriptional regulation and catalytic functions of enzymes could be the result of long-term evolutionary adaptation. The finding further expanded our understanding of GH-encoding genes amplification in filamentous fungi, which could guide the design of the optimal enzyme cocktails in industrial applications.

scholarly journals Peroxidase from Coleus Forskohlii: Purification and Biochemical Characterization

2020 ◽  
Vol 5 (1) ◽  
pp. 9-20
Author(s):  
Yaaser Q. Almulaiky ◽  
Yaaser Q. Almulaiky
Keyword(s):  
Ion Exchange ◽  
Peroxidase Activity ◽  
Biochemical Characterization ◽  
Specific Activity ◽  
Gel Filtration ◽  
Catalytic Efficiency ◽  
Industrial Applications ◽  
Coleus Forskohlii ◽  
Sds Page ◽  
Optimum Ph

In this study, a peroxidase from new source was purified using ion exchange and gel filtration techniques. The recovery for peroxidase activity was 19% with 11-fold purification and specific activity of 749 unit/mg protein. Purified peroxidase demonstrated a molecular mass of 39 kDa using gel filtration and was confirmed as a single band on SDS-PAGE. The purified peroxidase revealed a broad optimum pH activity at 6.0-6.5 and 50°C temperature. The kinetic parameters for purified peroxidase toward H2O2 and guaiacol as substrates were found to be Km = 3.355, 5.395 mM, Kcat = 99.52, 79.56 s-1 and Vmax =1.531, 1.242 µmole ml-1 min-1, respectively. The catalytic efficiency (kcat/Km) of the purified peroxidase was 14.75 and 29.66 s−1 mM−1 for guaiacol and H2O2, respectively. Peroxidase activity was observed to be enhanced by Cu2+, Co2+, Ni2+ and inhibited in the presence of Sn2+, Al3+, Hg2+, NaN3, EDTA and urea. Characterization showed that peroxidase purified from C. forskohlii has the ability to be used for food industrial applications.

Microbial keratinases: An overview of biochemical characterization and its eco-friendly approach for industrial applications

2020 ◽  
Vol 252 ◽  
pp. 119847 ◽  
Author(s):  
Binti Srivastava ◽  
Madhu Khatri ◽  
Gursharan Singh ◽  
Shailendra Kumar Arya
Keyword(s):  
Biochemical Characterization ◽  
Industrial Applications
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