Characterization of termite Trinervitermes trinervoides metagenome-derived glycoside hydrolases, the formulation of synergistic core enzyme sets for effective sweet sorghum and corncob saccharification, and their potential industrial applications

2019 ◽  
Author(s):  
◽  
Mpho Stephen Mafa
Keyword(s):  
Protein Structure ◽  
Sweet Sorghum ◽  
Metal Ion ◽  
Industrial Applications ◽  
Biochemical Properties ◽  
Product Inhibition ◽  
Glycoside Hydrolases ◽  
Core Enzyme ◽  
Polymeric Substrates ◽  
Hydrolysis Of

The current study investigated the biochemical properties of endo-glucanase (GH5E), exo-glucanase (GH5D), xylanase (GH5H) and endo-glucanase/xylanase (GH45), derived from the hindgut bacterial symbionts of a termite (Trinervitermes trinervoides) for their potential role in the biotechnology industry. All these enzymes, except GH5D, exhibited activities on cellulosic and xylan-rich polymeric substrates, which only displayed activity on p-nitrophenyl cellobioside. GH5D, GH5E, GH5H and GH45 enzymes retained more than 80% of their activities at pH 5.5 and also retained more than 80% of their activities at 40ºC. Furthermore, these enzymes were thermostable at 37ºC for 72 hours. GH5E, GH5H and GH45 were generally stable over a range of metal-ion. The kinetic parameters for GH5E were 5.68 mg/ml (KM) and 34.36 U/mg protein (Vmax). GH5D activity did not follow classical Michaelis-Menten kinetics, suggesting product inhibition. GH5H displayed KM values of 5.53, 95.03 and 2.10 mg/ml and Vmax values of 112.36, 144.45 and 180.32 U/mg protein on beechwood xylan, CMC, and xyloglucan, respectively. GH45 displayed a KM of 6.94 mg/ml and a Vmax of 12.30 U/mg protein on CMC. GH5D [cellobiohydrolase (CBH)] and a commercial CBHII (GH6) enzyme outperformed a commercial CBHI (GH7) enzyme when these enzymes hydrolysed β-glucan. GH5D and CBHII also displayed a higher degree of synergy on β-glucan but failed to show synergy on Avicel. We therefore concluded that GH5D and CBHII are β-glucan-specific cellobiohydrolases. The corncob (CC) and sweet sorghum bagasse (SSB) substrates were pretreated with lime, NaOH and NaClO2. Subsequent to pretreatment, these substrates were used to investigate if GH5D, GH5E, GH5H and GH45 could operate in synergy. Results revealed that out of 12 possible core enzyme sets constructed, only two (referred to as CES-E and CES-H) displayed higher activities on pretreated CC or SSB. Simultaneous synergy was generally the most effective mode of synergy during hydrolysis of alkaline pretreated SSB and CC samples by both CES-E and CES-H. Both core enzyme sets did not display synergy on oxidative pretreated substrates. These findings suggest that lime and NaOH are more effective pretreatments for CC and SSB substrates. We used PRotein Interactive MOdeling (PRIMO) software to demonstrate that GH5D protein structure is an (α/β)8 barrel with a tunnel-like active site. Enzymes with this type of protein structure are able to perform transglycosylation, a process in which GH5D produced methyl, ethyl and propyl cellobiosides. We concluded that the GH5D, GH5E, GH5H and GH45 enzymes possess novel biochemical properties and that they form synergy during the hydrolysis of complex substrates (SSB and CC). GH5D transglycosylation could be used to produce novel biodegradable chemicals with special properties (e.g. anti-microbial properties). In conclusion, our findings suggest that GH5D, GH5E, GH5H and GH45 can potentially be used to improve biorefinery processes.

scholarly journals Identification and molecular characterization of a psychrophilic GH1 β-glucosidase from the subtropical soil microorganism Exiguobacterium sp. GXG2

AMB Express ◽  
2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Bangqiao Yin ◽  
Hengsen Gu ◽  
Xueyan Mo ◽  
Yue Xu ◽  
Bing Yan ◽  
...  
Keyword(s):  
Metal Ion ◽  
Functional Characterization ◽  
Industrial Applications ◽  
Biochemical Properties ◽  
Screening Strategy ◽  
Soil Microorganism ◽  
Potential Candidate ◽  
Cold Adapted ◽  
Subtropical Soil

Abstract The products of bacterial β-glucosidases with favorable cold-adapted properties have industrial applications. A psychrophilic β-glucosidase gene named bglG from subtropical soil microorganism Exiguobacterium sp. GXG2 was isolated and characterized by function-based screening strategy. Results of multiple alignments showed that the derived protein BglG shared 45.7% identities with reviewed β-glucosidases in the UniProtKB/Swiss-Prot database. Functional characterization of the β-glucosidase BglG indicated that BglG was a 468 aa protein with a molecular weight of 53.2 kDa. The BglG showed the highest activity in pH 7.0 at 35 °C and exhibited consistently high levels of activity within low temperatures ranging from 5 to 35 °C. The BglG appeared to be a psychrophilic enzyme. The values of Km, Vmax, kcat, and kcat/Km of recombinant BglG toward ρNPG were 1.1 mM, 1.4 µg/mL/min, 12.7 s−1, and 11.5 mM/s, respectively. The specific enzyme activity of BglG was 12.14 U/mg. The metal ion of Ca2+ and Fe3+ could stimulate the activity of BglG, whereas Mn2+ inhibited the activity. The cold-adapted β-glucosidase BglG displayed remarkable biochemical properties, making it a potential candidate for future industrial applications.

scholarly journals A corynecaterium glutamicum expression system for glycoside hydrolase analysis

2021 ◽  
Author(s):  
Hirak Saxena
Keyword(s):  
Expression System ◽  
Lysosomal Storage Diseases ◽  
Industrial Applications ◽  
Glycoside Hydrolases ◽  
Cellulosic Biomass ◽  
Lysosomal Storage ◽  
Cellular Functions ◽  
Domains Of Life ◽  
Hydrolysis Of ◽  
Structure And Activity

The biological hydrolysis of glycosidic linkages in complex sugars is facilitated by glycoside hydrolases. These enzymes are ubiquitous across all domains of life, playing significant roles in important biological processes like the degradation of cellulosic biomass, viral pathogenesis, antibacterial defense, and normal cellular functions. The potential industrial applications of highly efficient glycoside hydrolases, as well as the fact that a number of lysosomal storage diseases have been attributed to deficiencies in these enzymes 43, 22, merits further study into their structure and activity. For this reason, a handful of novel glycoside hydrolases from Cellulomonas fimi, a Gram-positive Actinobacteria known for its ability to degrade cellulose 39, will be cloned, expressed and biochemically analyzed.

scholarly journals A corynecaterium glutamicum expression system for glycoside hydrolase analysis

2021 ◽  
Author(s):  
Hirak Saxena
Keyword(s):  
Expression System ◽  
Lysosomal Storage Diseases ◽  
Industrial Applications ◽  
Glycoside Hydrolases ◽  
Cellulosic Biomass ◽  
Lysosomal Storage ◽  
Cellular Functions ◽  
Domains Of Life ◽  
Hydrolysis Of ◽  
Structure And Activity

The biological hydrolysis of glycosidic linkages in complex sugars is facilitated by glycoside hydrolases. These enzymes are ubiquitous across all domains of life, playing significant roles in important biological processes like the degradation of cellulosic biomass, viral pathogenesis, antibacterial defense, and normal cellular functions. The potential industrial applications of highly efficient glycoside hydrolases, as well as the fact that a number of lysosomal storage diseases have been attributed to deficiencies in these enzymes 43, 22, merits further study into their structure and activity. For this reason, a handful of novel glycoside hydrolases from Cellulomonas fimi, a Gram-positive Actinobacteria known for its ability to degrade cellulose 39, will be cloned, expressed and biochemically analyzed.

scholarly journals Xyloglucan Oligosaccharides Hydrolysis by Exo-Acting Glycoside Hydrolases from Hyperthermophilic Microorganism Saccharolobus solfataricus

2021 ◽  
Vol 22 (7) ◽  
pp. 3325
Author(s):  
Nicola Curci ◽  
Andrea Strazzulli ◽  
Roberta Iacono ◽  
Federica De De Lise ◽  
Luisa Maurelli ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Industrial Applications ◽  
Apple Pomace ◽  
Glycoside Hydrolases ◽  
Hyperthermophilic Archaea ◽  
Industrial Processes ◽  
Degrading Enzymes ◽  
Hyperthermophilic Microorganism ◽  
Hydrolysis Of ◽  
Harsh Conditions

In the field of biocatalysis and the development of a bio-based economy, hemicellulases have attracted great interest for various applications in industrial processes. However, the study of the catalytic activity of the lignocellulose-degrading enzymes needs to be improved to achieve the efficient hydrolysis of plant biomasses. In this framework, hemicellulases from hyperthermophilic archaea show interesting features as biocatalysts and provide many advantages in industrial applications thanks to their stability in the harsh conditions encountered during the pretreatment process. However, the hemicellulases from archaea are less studied compared to their bacterial counterpart, and the activity of most of them has been barely tested on natural substrates. Here, we investigated the hydrolysis of xyloglucan oligosaccharides from two different plants by using, both synergistically and individually, three glycoside hydrolases from Saccharolobus solfataricus: a GH1 β-gluco-/β-galactosidase, a α-fucosidase belonging to GH29, and a α-xylosidase from GH31. The results showed that the three enzymes were able to release monosaccharides from xyloglucan oligosaccharides after incubation at 65 °C. The concerted actions of β-gluco-/β-galactosidase and the α-xylosidase on both xyloglucan oligosaccharides have been observed, while the α-fucosidase was capable of releasing all α-linked fucose units from xyloglucan from apple pomace, representing the first GH29 enzyme belonging to subfamily A that is active on xyloglucan.

scholarly journals BECLIN1: Protein Structure, Function and Regulation

Cells ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1522
Author(s):  
Sharon Tran ◽  
W. Douglas Fairlie ◽  
Erinna F. Lee
Keyword(s):  
Protein Structure ◽  
Structure Function ◽  
Membrane Trafficking ◽  
Biochemical Properties ◽  
Class Iii ◽  
Form Class ◽  
Protein Interactome ◽  
Protein Functions ◽  
Phosphoinositide 3 Kinase ◽  
Shed Light

BECLIN1 is a well-established regulator of autophagy, a process essential for mammalian survival. It functions in conjunction with other proteins to form Class III Phosphoinositide 3-Kinase (PI3K) complexes to generate phosphorylated phosphatidylinositol (PtdIns), lipids essential for not only autophagy but other membrane trafficking processes. Over the years, studies have elucidated the structural, biophysical, and biochemical properties of BECLIN1, which have shed light on how this protein functions to allosterically regulate these critical processes of autophagy and membrane trafficking. Here, we review these findings and how BECLIN1’s diverse protein interactome regulates it, as well as its impact on organismal physiology.

scholarly journals Carboxylic Ester Hydrolases in Bacteria: Active Site, Structure, Function and Application

Crystals ◽  
2019 ◽  
Vol 9 (11) ◽  
pp. 597 ◽  
Author(s):  
Changsuk Oh ◽  
T. Doohun Kim ◽  
Kyeong Kyu Kim
Keyword(s):  
Acyl Chain ◽  
Data Bank ◽  
Industrial Applications ◽  
Head Group ◽  
Carboxylic Ester ◽  
Site Structure ◽  
Domains Of Life ◽  
Carboxylic Esters ◽  
Hydrolysis Of ◽  
Ester Hydrolases

Carboxylic ester hydrolases (CEHs), which catalyze the hydrolysis of carboxylic esters to produce alcohol and acid, are identified in three domains of life. In the Protein Data Bank (PDB), 136 crystal structures of bacterial CEHs (424 PDB codes) from 52 genera and metagenome have been reported. In this review, we categorize these structures based on catalytic machinery, structure and substrate specificity to provide a comprehensive understanding of the bacterial CEHs. CEHs use Ser, Asp or water as a nucleophile to drive diverse catalytic machinery. The α/β/α sandwich architecture is most frequently found in CEHs, but 3-solenoid, β-barrel, up-down bundle, α/β/β/α 4-layer sandwich, 6 or 7 propeller and α/β barrel architectures are also found in these CEHs. Most are substrate-specific to various esters with types of head group and lengths of the acyl chain, but some CEHs exhibit peptidase or lactamase activities. CEHs are widely used in industrial applications, and are the objects of research in structure- or mutation-based protein engineering. Structural studies of CEHs are still necessary for understanding their biological roles, identifying their structure-based functions and structure-based engineering and their potential industrial applications.

scholarly journals “Newton’s cradle” proton relay with amide–imidic acid tautomerization in inverting cellulase visualized by neutron crystallography

2015 ◽  
Vol 1 (7) ◽  
pp. e1500263 ◽  
Author(s):  
Akihiko Nakamura ◽  
Takuya Ishida ◽  
Katsuhiro Kusaka ◽  
Taro Yamada ◽  
Shinya Fushinobu ◽  
...  
Keyword(s):  
Glycoside Hydrolases ◽  
Side Chain ◽  
Enzymatic Reactions ◽  
X Ray Diffraction ◽  
Peptide Bonds ◽  
Proton Relay ◽  
Newton’S Cradle ◽  
Dynamics Simulations ◽  
Hydrolysis Of

Hydrolysis of carbohydrates is a major bioreaction in nature, catalyzed by glycoside hydrolases (GHs). We used neutron diffraction and high-resolution x-ray diffraction analyses to investigate the hydrogen bond network in inverting cellulase PcCel45A, which is an endoglucanase belonging to subfamily C of GH family 45, isolated from the basidiomycete Phanerochaete chrysosporium. Examination of the enzyme and enzyme-ligand structures indicates a key role of multiple tautomerizations of asparagine residues and peptide bonds, which are finally connected to the other catalytic residue via typical side-chain hydrogen bonds, in forming the “Newton’s cradle”–like proton relay pathway of the catalytic cycle. Amide–imidic acid tautomerization of asparagine has not been taken into account in recent molecular dynamics simulations of not only cellulases but also general enzyme catalysis, and it may be necessary to reconsider our interpretation of many enzymatic reactions.

Metal Ion Catalysis in the Hydrolysis of Potassium Ethyl Oxalate

Nature ◽  
1964 ◽  
Vol 204 (4964) ◽  
pp. 1189-1190
Author(s):  
ROBERT W. HAY ◽  
NEIL J. WALKER
Keyword(s):  
Metal Ion ◽  
Metal Ion Catalysis ◽  
Hydrolysis Of

scholarly journals Metabolic engineering strategy for synthetizing trans-4-hydroxy-l-proline in microorganisms

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Zhenyu Zhang ◽  
Pengfu Liu ◽  
Weike Su ◽  
Huawei Zhang ◽  
Wenqian Xu ◽  
...  
Keyword(s):  
Metabolic Engineering ◽  
Biosynthetic Pathway ◽  
Industrial Applications ◽  
Microbial Cell Factories ◽  
Important Amino Acid ◽  
Cell Factories ◽  
Complex Process ◽  
Engineering Strategy ◽  
Hydrolysis Of ◽  
New Strategies

AbstractTrans-4-hydroxy-l-proline is an important amino acid that is widely used in medicinal and industrial applications, particularly as a valuable chiral building block for the organic synthesis of pharmaceuticals. Traditionally, trans-4-hydroxy-l-proline is produced by the acidic hydrolysis of collagen, but this process has serious drawbacks, such as low productivity, a complex process and heavy environmental pollution. Presently, trans-4-hydroxy-l-proline is mainly produced via fermentative production by microorganisms. Some recently published advances in metabolic engineering have been used to effectively construct microbial cell factories that have improved the trans-4-hydroxy-l-proline biosynthetic pathway. To probe the potential of microorganisms for trans-4-hydroxy-l-proline production, new strategies and tools must be proposed. In this review, we provide a comprehensive understanding of trans-4-hydroxy-l-proline, including its biosynthetic pathway, proline hydroxylases and production by metabolic engineering, with a focus on improving its production.

Sign in / Sign up

Export Citation Format

Share Document