scholarly journals Comparative Analysis and Biochemical Characterization of Two Endo-β-1,3-Glucanases from the Thermophilic Bacterium Fervidobacterium sp.

Catalysts ◽  
2019 ◽  
Vol 9 (10) ◽  
pp. 830 ◽  
Author(s):  
Burkhardt ◽  
Schäfers ◽  
Claren ◽  
Schirrmacher ◽  
Antranikian
Keyword(s):  
Elevated Temperatures ◽  
Biochemical Characterization ◽  
Hot Spring ◽  
Biomass Conversion ◽  
Catalytic Efficiency ◽  
Industrial Applications ◽  
Thermophilic Bacterium ◽  
Metagenomic Sample ◽  
Wide Range ◽  
Active Site Cleft

Laminarinases exhibit potential in a wide range of industrial applications including the production of biofuels and pharmaceuticals. In this study, we present the genetic and biochemical characteristics of FLamA and FLamB, two laminarinases derived from a metagenomic sample from a hot spring in the Azores. Sequence comparison revealed that both genes had high similarities to genes from Fervidobacterium nodosum Rt17-B1. The two proteins showed sequence similarities of 62% to each other and belong to the glycoside hydrolase (GH) family 16. For biochemical characterization, both laminarinases were heterologously produced in Escherichia coli and purified to homogeneity. FLamA and FLamB exhibited similar properties and both showed highest activity towards laminarin at 90 °C and pH 6.5. The two enzymes were thermostable but differed in their half-life at 80 °C with 5 h and 1 h for FLamA and FLamB, respectively. In contrast to other laminarinases, both enzymes prefer β-1,3-glucans and mixed-linked glucans as substrates. However, FLamA and FLamB differ in their catalytic efficiency towards laminarin. Structure predictions were made and showed minor differences particularly in a kink adjacent to the active site cleft. The high specific activities and resistance to elevated temperatures and various additives make both enzymes suitable candidates for application in biomass conversion.

scholarly journals Insights into Brevibacillus borstelensis AK1 through Whole Genome Sequencing: A Thermophilic Bacterium Isolated from a Hot Spring in Saudi Arabia

2018 ◽  
Vol 2018 ◽  
pp. 1-9 ◽  
Author(s):  
Amjad B. Khalil ◽  
Neelamegam Sivakumar ◽  
Muhammad Arslan ◽  
Hamna Saleem ◽  
Sami Qarawi
Keyword(s):  
Saudi Arabia ◽  
Hot Spring ◽  
Gc Content ◽  
Morphological Characterization ◽  
Thermophilic Bacterium ◽  
Whole Genome Sequence ◽  
Strictly Aerobic ◽  
Whole Genome ◽  
Aerobic Bacterium ◽  
Wide Range

Brevibacillus borstelensis AK1 is a thermophile which grows between the temperatures of 45°C and 70°C. The present study is an extended genome report of B. borstelensis AK1 along with the morphological characterization. The strain is isolated from a hot spring in Saudi Arabia (southeast of the city Gazan). It is observed that the strain AK1 is rod-shaped, motile, and strictly aerobic bacterium. The whole genome sequence resulted in 29 contigs with a total length of 5,155,092 bp. In total, 3,946 protein-coding genes and 139 RNA genes were identified. Comparison with the previously submitted strains of B. borstelensis strains illustrates that strain AK1 has a small genome size but high GC content. The strain possesses putative genes for degradation of a wide range of substrates including polyethylene (plastic) and long-chain hydrocarbons. These genomic features may be useful for future environmental/biotechnological applications.

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 Dye Decoloring Peroxidase Structure, Catalytic Properties and Applications: Current Advancement and Futurity

Catalysts ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 955
Author(s):  
Lingxia Xu ◽  
Jianzhong Sun ◽  
Majjid A. Qaria ◽  
Lu Gao ◽  
Daochen Zhu
Keyword(s):  
Protein Structure ◽  
Amino Acid ◽  
High Efficiency ◽  
Chemical Properties ◽  
Catalytic Efficiency ◽  
Industrial Applications ◽  
Research Strategy ◽  
Amino Acid Sequences ◽  
Alkali Resistance ◽  
Wide Range

Dye decoloring peroxidases (DyPs) were named after their high efficiency to decolorize and degrade a wide range of dyes. DyPs are a type of heme peroxidase and are quite different from known heme peroxidases in terms of amino acid sequences, protein structure, catalytic residues, and physical and chemical properties. DyPs oxidize polycyclic dyes and phenolic compounds. Thus they find high application potentials in dealing with environmental problems. The structure and catalytic characteristics of DyPs of different families from the amino acid sequence, protein structure, and enzymatic properties, and analyzes the high-efficiency degradation ability of some DyPs in dye and lignin degradation, which vary greatly among DyPs classes. In addition, application prospects of DyPs in biomedicine and other fields are also discussed briefly. At the same time, the research strategy based on genetic engineering and synthetic biology in improving the stability and catalytic activity of DyPs are summarized along with the important industrial applications of DyPs and associated challenges. Moreover, according to the current research findings, bringing DyPs to the industrial level may require improving the catalytic efficiency of DyP, increasing production, and enhancing alkali resistance and toxicity.

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.

scholarly journals Complex Electrical Conductivity of Biotite and Muscovite Micas at Elevated Temperatures: A Comparative Study

Materials ◽  
2020 ◽  
Vol 13 (16) ◽  
pp. 3513
Author(s):  
Vassilios Saltas ◽  
Despoina Pentari ◽  
Filippos Vallianatos
Keyword(s):  
Electrical Conductivity ◽  
Elevated Temperatures ◽  
Small Polaron ◽  
Complex Impedance ◽  
Bound Water ◽  
Data Representation ◽  
Industrial Applications ◽  
Mechanical And Thermal Properties ◽  
Wide Range ◽  
Complex Electrical Conductivity

The unique physicochemical, electrical, mechanical, and thermal properties of micas make them suitable for a wide range of industrial applications, and thus, the interest for these kind of hydrous aluminosilicate minerals is still persistent, not only from a practical but also from a scientific point of view. In the present work, complex impedance spectroscopy measurements were carried out in muscovite and biotite micas, perpendicular to their cleavage planes, over a broad range of frequencies (10−2 Hz to 106 Hz) and temperatures (473–1173 K) that have not been measured so far. Different formalisms of data representation were used, namely, Cole-Cole plots of complex impedance, complex electrical conductivity and electric modulus to analyze the electrical behavior of micas and the electrical signatures of the dehydration/dehydroxylation processes. Our results suggest that ac-conductivity is affected by the structural hydroxyls and the different concentrations of transition metals (Fe, Ti and Mg) in biotite and muscovite micas. The estimated activation energies, i.e., 0.33–0.83 eV for biotite and 0.69–1.92 eV for muscovite, were attributed to proton and small polaron conduction, due to the bound water and different oxidation states of Fe.

scholarly journals A thermostable Cas9 with increased lifetime in human plasma

2017 ◽  
Author(s):  
Lucas B. Harrington ◽  
David Paez-Espino ◽  
Janice S. Chen ◽  
Enbo Ma ◽  
Brett T. Staahl ◽  
...  
Keyword(s):  
Human Plasma ◽  
Dna Cleavage ◽  
Elevated Temperatures ◽  
Thermophilic Bacterium ◽  
Temperature Range ◽  
Cas9 Protein ◽  
Wide Range ◽  
Rnp Complex ◽  
Mammalian Genomes

CRISPR-Cas9 is a powerful technology that has enabled genome editing in a wide range of species. However, the currently developed Cas9 homologs all originate from mesophilic bacteria, making them susceptible to proteolytic degradation and unsuitable for applications requiring function at elevated temperatures. Here, we show that the Cas9 protein from the thermophilic bacteriumGeobacillus stearothermophilus(GeoCas9) catalyzes RNA-guided DNA cleavage over a wide temperature range and has an enhanced protein lifetime in human plasma. GeoCas9 is active at temperatures up to 70°C, compared to 45°C forStreptococcus pyogenesCas9 (SpyCas9), which greatly expands the temperature range for CRISPR-Cas9 applications. By comparing features of two closely relatedGeobacillushomologs, we created a variant of GeoCas9 that doubles the DNA target sequences that can be recognized by this system. We also found that GeoCas9 is an effective tool for editing mammalian genomes when delivered as a ribonucleoprotein (RNP) complex. Together with an increased lifetime in human plasma, the thermostable GeoCas9 provides the foundation for improved RNP deliveryin vivoand expands the temperature range of CRISPR-Cas9.

scholarly journals CFD Modeling of Gas–Solid Cyclone Separators at Ambient and Elevated Temperatures

Processes ◽  
2020 ◽  
Vol 8 (2) ◽  
pp. 228 ◽  
Author(s):  
Mohammadhadi Nakhaei ◽  
Bona Lu ◽  
Yujie Tian ◽  
Wei Wang ◽  
Kim Dam-Johansen ◽  
...  
Keyword(s):  
Elevated Temperatures ◽  
Cfd Simulation ◽  
Separation Efficiency ◽  
Flow Behavior ◽  
Industrial Applications ◽  
High Mass ◽  
Cfd Modeling ◽  
Solid Flow ◽  
Solids Loading ◽  
Wide Range

Gas–solid cyclone separators are widely utilized in many industrial applications and usually involve complex multi-physics of gas–solid flow and heat transfer. In recent years, there has been a progressive interest in the application of computational fluid dynamics (CFD) to understand the gas–solid flow behavior of cyclones and predict their performance. In this paper, a review of the existing CFD studies of cyclone separators, operating in a wide range of solids loadings and at ambient and elevated temperatures, is presented. In the first part, a brief background on the important performance parameters of cyclones, namely pressure drop and separation efficiency, as well as how they are affected by the solids loading and operating temperature, is described. This is followed by a summary of the existing CFD simulation studies of cyclones at ambient temperature, with an emphasis on the high mass loading of particles, and at elevated temperatures. The capabilities as well as the challenges and limitations of the existing CFD approaches in predicting the performance of cyclones operating in such conditions are evaluated. Finally, an outlook on the prospects of CFD simulation of cyclone separators is provided.

scholarly journals Programmable DNA cleavage by Ago nucleases from mesophilic bacteria Clostridium butyricum and Limnothrix rosea

2019 ◽  
Author(s):  
Anton Kuzmenko ◽  
Denis Yudin ◽  
Sergei Ryazansky ◽  
Andrey Kulbachinskiy ◽  
Alexei A. Aravin
Keyword(s):  
Dna Cleavage ◽  
Elevated Temperatures ◽  
Biochemical Characterization ◽  
Clostridium Butyricum ◽  
Seed Region ◽  
Mesophilic Bacteria ◽  
Highly Active ◽  
Wide Range

ABSTRACTArgonaute (Ago) proteins are the key players in RNA interference in eukaryotes, where they function as RNA-guided RNA endonucleases. Prokaryotic Argonautes (pAgos) are much more diverse than their eukaryotic counterparts but their cellular functions and mechanisms of action remain largely unknown. Some pAgos were shown to use small DNA guides for endonucleolytic cleave of complementary DNA in vitro. However, previously studied pAgos from thermophilic prokaryotes function at elevated temperatures which limits their potential use as a tool in genomic applications. Here, we describe two pAgos from mesophilic bacteria, Clostridium butyricum (CbAgo) and Limnothrix rosea (LrAgo), that act as DNA-guided DNA nucleases at physiological temperatures. In contrast to previously studied pAgos, CbAgo and LrAgo can use not only 5’-phosphorylated but also 5’-hydroxyl DNA guides, with diminished precision of target cleavage. Both LrAgo and CbAgo can tolerate guide/target mismatches in the seed region, but are sensitive to mismatches in the 3’-guide region. CbAgo is highly active under a wide range of conditions and can be used for programmable endonucleolytic cleavage of both single-stranded and double-stranded DNA substrates at moderate temperatures. The biochemical characterization of mesophilic pAgo proteins paths the way for their use for DNA manipulations both in vitro and in vivo.

scholarly journals Statistical factorial designs for optimum production of thermostable α-amylase by the degradative bacterium Parageobacillus thermoglucosidasius Pharon1 isolated from Sinai, Egypt

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Ali M. Saeed ◽  
Einas H. El-Shatoury ◽  
Hayam A. E. Sayed
Keyword(s):  
Response Surface ◽  
Hot Spring ◽  
16S Rrna Gene Sequencing ◽  
Soluble Starch ◽  
Industrial Applications ◽  
Thermophilic Bacterium ◽  
Rrna Gene ◽  
Amylase Production ◽  
Full Factorial ◽  
Parageobacillus Thermoglucosidasius

Abstract Background This study aimed to isolate potent thermophilic and amylolytic bacteria from a hot spring of Pharaoh’s bath, Sinai, Egypt, and screen its degradative activity. The amylolytic activity was further optimized using a statistical full factorial design followed by response surface methodology. Results A thermophilic bacterium was isolated from the hot spring of Pharaoh’s Bath, Sinai, Egypt. The isolate produced amylase, cellulase, and caseinase and was identified by 16S rRNA gene sequencing as Parageobacillus thermoglucosidasius Pharon1 (MG965879). A growth medium containing 1% soluble starch was found to optimize the amylase production. Dinitrosalycalic acid method (DNS) was used to estimate the amount of reducing sugar produced. Statistical full factorial and response surface designs were employed to optimize physical variables affecting the α-amylase production and determine the significant interactions of the studied variables during the fermentation process. According to the results obtained by the response optimizer, the maximum amylase activity reached 76.07 U/mL/ min at 54.1°C, pH 5.6 after 98.5 h incubation under aerobic conditions. Moreover, the produced enzyme was thermostable and retained most of its activity when exposed to a high temperature of 100°C for 120 min. Maximum enzyme activity was attained when the enzyme was incubated at 70°C for 30 min. Conclusions This is the first report of the production of thermostable α-amylase by the potent thermophilic Parageobacillus thermoglucosidasius. The enzyme endured extreme conditions of temperature and pH which are important criteria for commercial and industrial applications.

scholarly journals Novel halo- and thermo-tolerant Cohnella sp. A01 L-glutaminase: heterologous expression and biochemical characterization

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Samaneh Mosallatpour ◽  
Saeed Aminzadeh ◽  
Mehdi Shamsara ◽  
Reza Hajihosseini
Keyword(s):  
Food Industry ◽  
Tertiary Structure ◽  
Biochemical Characterization ◽  
Arrhenius Plot ◽  
Specific Activity ◽  
Catalytic Efficiency ◽  
Cysteine Residue ◽  
Enzyme Active Site ◽  
Wide Range ◽  
Ph Range

AbstractL-glutaminase importance to use in the food industry and medicine has attracted much attention. Enzymes stability has always been a challenge while working with them. We heterologously expressed and characterized a novel stable L-glutaminase from an extremophile bacterium (Cohnella sp. A01, PTCC No: 1921). Km, Vmax, catalytic efficiency and specific activity of rSAM were respectively 1.8 mM, 49 µmol/min, 1851 1/(S.mM) and 9.2 IU/mg. Activation energy for substrate to product conversion and irreversible thermo-inactivation were respectively 4 kJ/mol and 105 kJ/mol from the linear Arrhenius plot. rSAM had the highest activity at temperature 50 °C, pH 8 and was resistant to a wide range of temperature and pH. In compare to the other characterized glutaminases, rSAM was the most resistant to NaCl. Mg2+, glycerol, DTT, and BME enhanced the enzyme activity and iodoacetate and iodoacetamide inhibited it. rSAM had only been partially digested by some proteases. According to the Fluorimetry and Circular dichroism analysis, rSAM in pH range from 4 to 11 and temperatures up to 60 °C had structural stability. A cysteine residue in the enzyme active site and a thiol bond were predicted upon the modeled tertiary structure of rSAM. Present structural studies also confirmed the presence of a thiol bond in its structure.

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