scholarly journals Archaeal and bacterial thermostable amylopullulanases: characteristic features and biotechnological applications

Amylase ◽  
2018 ◽  
Vol 2 (1) ◽  
pp. 44-57
Author(s):  
Tulasi Satyanarayana ◽  
Mohanan Nisha
Keyword(s):  
Catalytic Domain ◽  
Bread Making ◽  
Amylolytic Enzymes ◽  
Conserved Sequence ◽  
Glycosidic Bonds ◽  
Bifunctional Enzymes ◽  
One Step ◽  
Starch Saccharification ◽  
Characteristic Features ◽  
The Cost

AbstractAmylopullulanases are endoacting bifunctional enzymes capable of hydrolyzing α-1,4- and α-1,6-glycosidic linkages in starch, amylose, pullulan, amylopectin and related oligosaccharides. These enzymes possess single or dual active site(s) for cleaving α-1,4- and α-1,6-glycosidic bonds; the former are called amylopullulanases, and the latter, α-amylase-pullulanases. These are grouped into GH13 and GH57 families based on the architecture of the catalytic domain and the number of conserved sequence regions. The amylopullulanases/α-amylasepullulanases are produced by bacteria as well as archaea, and among them, thermophilic and hyperthermophilic species are the major producers. The thermostable amylopullulanases find application in one-step starch liquefaction-saccharification to form various sugar syrups and maltooligosaccharides. The starch saccharification process catalysed by amylopullulanases minimizes the use of other amylolytic enzymes, like α-amylase and glucoamylase, thereby reducing the cost of sugar syrups. The enzymes also find applications in bread making as an anti-stale and as a detergent additive.

scholarly journals Amylolytic enzymes - focus on the alpha-amylases from Archae and plants

2021 ◽  
Vol 9 (1) ◽  
pp. 5-26
Author(s):  
Štefan Janeček
Keyword(s):  
Three Dimensional ◽  
Structural Features ◽  
Catalytic Triad ◽  
Amino Acid Sequences ◽  
Glycoside Hydrolases ◽  
Amylolytic Enzymes ◽  
Conserved Sequence ◽  
Glycosidic Bonds ◽  
Evolutionary Relatedness ◽  
Tim Barrel

Amylolytic enzymes represent a group of starch hydrolases and related enzymes that are active towards the α-glycosidic bonds in starch and related poly- and oligosaccharides. The three best known amylolytic enzymes are α-amylase, β-amylase and glucoamylase that, however, differ from each other by their amino acid sequences, three-dimensional structures, reaction mechanisms and catalytic machineries. In the sequence-based classification of all glycoside hydrolases (GHs) they have therefore been classified into the three independent families: GH13 (α-amylases), GH14 (β-amylases) and GH15 (glucoamylases). Some amylolytic enzymes have been placed to the families GH31 and GH57. The family GH13 together with the families GH70 and GH77 constitutes the clan GH-H, well-known as the α-amylase family. It contains more than 6,000 sequences and covers 30 various enzyme specificities sharing the conserved sequence regions, catalytic TIM-barrel fold, retaining reaction mechanism and catalytic triad. Among the GH13 α-amylases, those produced by plants and archaebacteria exhibit common sequence similarities that distinguish them from the α-amylases of the remaining taxonomic sources. Despite the close evolutionary relatedness between the plant and archaeal α-amylases, there are also specific differences that discriminate them from each other. These specific differences could be used in an effort to reveal the sequence-structural features responsible for the high thermostability of the α-amylases from Archaea.

The first crystal structure of the peptidase domain of the U32 peptidase family

2015 ◽  
Vol 71 (12) ◽  
pp. 2505-2512 ◽  
Author(s):  
Magdalena Schacherl ◽  
Angelika A. M. Montada ◽  
Elena Brunstein ◽  
Ulrich Baumann
Keyword(s):  
Crystal Structure ◽  
Catalytic Mechanism ◽  
Quaternary Structure ◽  
Catalytic Domain ◽  
Three Dimensional ◽  
Zinc Ion ◽  
Crystal Structure Analysis ◽  
Dimensional Structure ◽  
Sequence Motifs ◽  
Conserved Sequence

The U32 family is a collection of over 2500 annotated peptidases in the MEROPS database with unknown catalytic mechanism. They mainly occur in bacteria and archaea, but a few representatives have also been identified in eukarya. Many of the U32 members have been linked to pathogenicity, such as proteins fromHelicobacterandSalmonella. The first crystal structure analysis of a U32 catalytic domain fromMethanopyrus kandleri(genemk0906) reveals a modified (βα)8TIM-barrel fold with some unique features. The connecting segment between strands β7 and β8 is extended and helix α7 is located on top of the C-terminal end of the barrel body. The protein exhibits a dimeric quaternary structure in which a zinc ion is symmetrically bound by histidine and cysteine side chains from both monomers. These residues reside in conserved sequence motifs. No typical proteolytic motifs are discernible in the three-dimensional structure, and biochemical assays failed to demonstrate proteolytic activity. A tunnel in which an acetate ion is bound is located in the C-terminal part of the β-barrel. Two hydrophobic grooves lead to a tunnel at the C-terminal end of the barrel in which an acetate ion is bound. One of the grooves binds to aStrep-Tag II of another dimer in the crystal lattice. Thus, these grooves may be binding sites for hydrophobic peptides or other ligands.

THE TECHNICAL CONDITION OF THE SURVEYED HYDRAULIC UNITS IN THE MOSCOW REGION AND THE CLASSIFICATION FEATURES OF THEIR RESERVOIRS

2021 ◽  
pp. 69-79
Author(s):  
V. V. GRITSAN ◽  
Keyword(s):  
Technical Condition ◽  
Moscow Region ◽  
Risk Level ◽  
Ecological Framework ◽  
Safety Level ◽  
Repair Work ◽  
Hydraulic Unit ◽  
Technical Parameters ◽  
Characteristic Features ◽  
The Cost

The article presents the results of surveys of 311 class IV hydraulic structures carried out in 2016-2020 in the Moscow region. All the reservoirs of the surveyed hydraulic units were classified according to their characteristic features, the technical condition of culverts and dams was assessed, there was established the safety level of both separate structures and hydraulic units as a whole. During the surveys, the technical parameters of the surveyed structures were established, the state of each structure and the hydraulic unit as a whole was assessed, a possibility of their accident and a risk level for the downstream areas were considered. At the same time, recommendations were developed for the elimination of serious damage and, with the help of an examination, the amount of the cost of the necessary repair work was determined. The paper also assesses the issues of the ecological state of the areas where the hydraulic units are located and the hydraulic units themselves as blocks of the ecological framework of the territories.

scholarly journals The Crystal Structure of Shikimate Dehydrogenase (AroE) Reveals a Unique NADPH Binding Mode

2003 ◽  
Vol 185 (14) ◽  
pp. 4144-4151 ◽  
Author(s):  
Sheng Ye ◽  
Frank von Delft ◽  
Alexei Brooun ◽  
Mark W. Knuth ◽  
Ronald V. Swanson ◽  
...  
Keyword(s):  
Catalytic Domain ◽  
Binding Pocket ◽  
Binding Mode ◽  
Sequence Motif ◽  
Binary Complex ◽  
Shikimate Dehydrogenase ◽  
Conserved Residues ◽  
Conserved Sequence ◽  
Nicotinamide Mononucleotide ◽  
Reversible Reduction

ABSTRACT Shikimate dehydrogenase catalyzes the NADPH-dependent reversible reduction of 3-dehydroshikimate to shikimate. We report the first X-ray structure of shikimate dehydrogenase from Haemophilus influenzae to 2.4-Å resolution and its complex with NADPH to 1.95-Å resolution. The molecule contains two domains, a catalytic domain with a novel open twisted α/β motif and an NADPH binding domain with a typical Rossmann fold. The enzyme contains a unique glycine-rich P-loop with a conserved sequence motif, GAGGXX, that results in NADPH adopting a nonstandard binding mode with the nicotinamide and ribose moieties disordered in the binary complex. A deep pocket with a narrow entrance between the two domains, containing strictly conserved residues primarily contributed by the catalytic domain, is identified as a potential 3-dehydroshikimate binding pocket. The flexibility of the nicotinamide mononucleotide portion of NADPH may be necessary for the substrate 3-dehydroshikimate to enter the pocket and for the release of the product shikimate.

scholarly journals Structural and Catalytic Characterization of TsBGL, a β-Glucosidase From Thermofilum sp. ex4484_79

2021 ◽  
Vol 12 ◽  
Author(s):  
Anke Chen ◽  
Dan Wang ◽  
Rui Ji ◽  
Jixi Li ◽  
Shaohua Gu ◽  
...  
Keyword(s):  
Catalytic Domain ◽  
Specific Activity ◽  
Maximum Activity ◽  
Homologous Proteins ◽  
Glycosidic Bonds ◽  
Catalytic Sites ◽  
Potential Applications ◽  
Beta Glucosidase ◽  
Hydrolysis Of

Beta-glucosidase is an enzyme that catalyzes the hydrolysis of the glycosidic bonds of cellobiose, resulting in the production of glucose, which is an important step for the effective utilization of cellulose. In the present study, a thermostable β-glucosidase was isolated and purified from the Thermoprotei Thermofilum sp. ex4484_79 and subjected to enzymatic and structural characterization. The purified β-glucosidase (TsBGL) exhibited maximum activity at 90°C and pH 5.0 and displayed maximum specific activity of 139.2μmol/min/mgzne against p-nitrophenyl β-D-glucopyranoside (pNPGlc) and 24.3μmol/min/mgzen against cellobiose. Furthermore, TsBGL exhibited a relatively high thermostability, retaining 84 and 47% of its activity after incubation at 85°C for 1.5h and 90°C for 1.5h, respectively. The crystal structure of TsBGL was resolved at a resolution of 2.14Å, which revealed a classical (α/β)8-barrel catalytic domain. A structural comparison of TsBGL with other homologous proteins revealed that its catalytic sites included Glu210 and Glu414. We provide the molecular structure of TsBGL and the possibility of improving its characteristics for potential applications in industries.

scholarly journals One-Step Bark-Like Imitated Polypropylene (PP)/Polycarbonate (PC) Nanofibrous Meltblown Membrane for Efficient Particulate Matter Removal

Polymers ◽  
2019 ◽  
Vol 11 (8) ◽  
pp. 1307 ◽  
Author(s):  
Ting-Ting Li ◽  
Xixi Cen ◽  
Hai-Tao Ren ◽  
Fei Sun ◽  
Qi Lin ◽  
...  
Keyword(s):  
Particulate Matter ◽  
Fiber Diameter ◽  
Average Fiber Diameter ◽  
Efficient Removal ◽  
Filtration Performance ◽  
Innovative Method ◽  
High Efficient ◽  
One Step ◽  
The Cost ◽  
Pm Removal

A bark-like imitated polypr opylene (PP)/polycarbonate (PC) nanofibrous membrane was constructed by one-step meltblown technique for efficient particulate matter (PM) removal. The effects of PC content (0%, 1%, 3%, 5%, and 7%) on membrane thermal stability, microscopic characteristics, filtration performance, hydrophilicity, and water vapor transmission were investigated. The results demonstrated that using facile design of incompatibility and viscosity difference between PC and PP polymers decreases average fiber diameter, creating a bark-like groove appearance and increasing surface potential, making a new PP/PC membrane with high filtration performance. The resultant PP/PC membrane had finer average fiber diameter of 0.63 μm, which was nearly 89.41% lower than PP membranes (5.95 μm), and its quality factor (0.036 Pa−1) was nearly 2.12 times than that of PP membranes (0.017 Pa−1) with the die hole diameter of 0.5 mm. This fabrication technique of a special meltblown filter membrane saves the cost of die retrofitting and post-processing, which provides an innovative method for particulate efficient removal of high efficient filters.

scholarly journals Glucoamylase starch-binding domain of Aspergillus niger B1: molecular cloning and functional characterization

2003 ◽  
Vol 372 (3) ◽  
pp. 905-910 ◽  
Author(s):  
Tzur PALDI ◽  
Ilan LEVY ◽  
Oded SHOSEYOV
Keyword(s):  
Aspergillus Niger ◽  
Corn Starch ◽  
Catalytic Domain ◽  
Functional Characterization ◽  
Carbohydrate Binding ◽  
Amylolytic Enzymes ◽  
Binding Domains ◽  
C Terminus ◽  
Carbohydrate Binding Modules ◽  
Modified Starches

Carbohydrate-binding modules (CBMs) are protein domains located within a carbohydrate-active enzyme, with a discrete fold that can be separated from the catalytic domain. Starch-binding domains (SBDs) are CBMs that are usually found at the C-terminus in many amylolytic enzymes. The SBD from Aspergillus niger B1 (CMI CC 324262) was cloned and expressed in Escherichia coli as an independent domain and the recombinant protein was purified on starch. The A. niger B1 SBD was found to be similar to SBD from A. kawachii, A. niger var. awamori and A. shirusami (95–96% identity) and was classified as a member of the CBM family 20. Characterization of SBD binding to starch indicated that it is essentially irreversible and that its affinity to cationic or anionic starch, as well as to potato or corn starch, does not differ significantly. These observations indicate that the fundamental binding area on these starches is essentially the same. Natural and chemically modified starches are among the most useful biopolymers employed in the industry. Our study demonstrates that SBD binds effectively to both anionic and cationic starch.

scholarly journals Identification of three critical acidic residues of poly(ADP-ribose) glycohydrolase involved in catalysis: determining the PARG catalytic domain

2005 ◽  
Vol 388 (2) ◽  
pp. 493-500 ◽  
Author(s):  
Chandra N. PATEL ◽  
David W. KOH ◽  
Myron K. JACOBSON ◽  
Marcos A. OLIVEIRA
Keyword(s):  
Catalytic Activity ◽  
Catalytic Domain ◽  
Functional Characterization ◽  
Sequence Alignments ◽  
Inhibitor Binding ◽  
Conserved Sequence ◽  
Binding Residue ◽  
Acidic Residues ◽  
Hydrolysis Of

PARG [poly(ADP-ribose) glycohydrolase] catalyses the hydrolysis of α(1″→2′) or α(1‴→2″) O-glycosidic linkages of ADP-ribose polymers to produce free ADP-ribose. We investigated possible mechanistic similarities between PARG and glycosidases, which also cleave O-glycosidic linkages. Glycosidases typically utilize two acidic residues for catalysis, thus we targeted acidic residues within a conserved region of bovine PARG that has been shown to contain an inhibitor-binding site. The targeted glutamate and aspartate residues were changed to asparagine in order to minimize structural alterations. Mutants were purified and assayed for catalytic activity, as well as binding, to an immobilized PARG inhibitor to determine ability to recognize substrate. Our investigation revealed residues essential for PARG catalytic activity. Two adjacent glutamic acid residues are found in the conserved sequence Gln755-Glu-Glu757, and a third residue found in the conserved sequence Val737-Asp-Phe-Ala-Asn741. Our functional characterization of PARG residues, along with recent identification of an inhibitor-binding residue Tyr796 and a glycine-rich region Gly745-Gly-Gly747 important for PARG function, allowed us to define a PARG ‘signature sequence’ [vDFA-X3-GGg-X6–8-vQEEIRF-X3-PE-X14-E-X12-YTGYa], which we used to identify putative PARG sequences across a range of organisms. Sequence alignments, along with our mapping of PARG functional residues, suggest the presence of a conserved catalytic domain of approx. 185 residues which spans residues 610–795 in bovine PARG.

scholarly journals Engineering of Bifunctional Enzymes with Uricase and Peroxidase Activities for Simple and Rapid Quantification of Uric Acid in Biological Samples

Catalysts ◽  
2020 ◽  
Vol 10 (4) ◽  
pp. 428
Author(s):  
Thanawat Phuadraksa ◽  
Jurairat Chittrakanwong ◽  
Kittitouch Tullayaprayouch ◽  
Naruthai Onsirisakul ◽  
Sineewanlaya Wichit ◽  
...  
Keyword(s):  
Uric Acid ◽  
Low Cost ◽  
Colorimetric Detection ◽  
Amplex Red ◽  
Bifunctional Enzymes ◽  
Bifunctional Proteins ◽  
Frame Fusion ◽  
The Cost ◽  
Optimized Conditions ◽  
Catalyzed Oxidation

Serum uric acid (SUA) is an important biomarker for prognosis and management of gout and other diseases. The development of a low-cost, simple, rapid and reliable assay for SUA detection is of great importance. In the present study, to save the cost of enzyme production and to shorten the reaction time for uric acid quantification, bifunctional proteins with uricase and peroxidase activities were engineered. In-frame fusion of Candida utilis uricase (CUOX) and Vitreoscilla hemoglobin (VHb) resulted in two versions of the bifunctional protein, CUOX-VHb (CV) and VHb-CUOX (VC). To our knowledge, this is the first report to describe the production of proteins with uricase and peroxidase activities. Based on the measurement of the initial rates of the coupled reaction (between uricase and peroxidase), CV was proven to be the most efficient enzyme followed by VC and native enzymes (CUOX+VHb), respectively. CV was further applied for the development of an assay for colorimetric detection of SUA, which was based on VHb-catalyzed oxidation of Amplex Red in the presence of hydrogen peroxide (H2O2). Under the optimized conditions, the assay exhibited a linear relationship between the absorbance and UA concentration over the range of 2.5 to 50 μM, with a detection limit of 1 μM. In addition, the assay can be performed at a single pH (8.0) so adjustment of the pH for peroxidase activity was not required. This advantage helped to further reduce costs and time. The developed assay was also successfully applied to detect UA in pooled human serum with the recoveries over 94.8%. These results suggest that the proposed assay holds great potential for clinical application.

Controllable fabrication of α-Ni(OH)2 thin films with preheating treatment for long-term stable electrochromic and energy storage applications

2020 ◽  
Vol 8 (9) ◽  
pp. 3010-3016 ◽  
Author(s):  
Chunhua Su ◽  
Meijia Qiu ◽  
Yipeng An ◽  
Siyuan Sun ◽  
Chuanxi Zhao ◽  
...  
Keyword(s):  
Thin Films ◽  
Energy Storage ◽  
Cost Effective ◽  
Fast Switching ◽  
Energy Storage Application ◽  
One Step ◽  
Energy Storage Applications ◽  
Controllable Fabrication ◽  
The Cost

Long-term stable, fast switching α-Ni(OH)2 electrodes for electrochromic energy storage application has been fabricated by one-step preheating treatment. The cost-effective fabrication is expected to expand to other metal hydroxide materials.

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