A single site mutation significantly improves the thermostability and activity of heparinase I from<i>Bacteroides eggerthii</i>

Su, Wen-Bin; Zhu, Chenyuan; Zhou, Hongbo; Gao, Jian; Zhang, Ye-Wang

doi:10.1080/10242422.2021.1976757

Cited by 3 publications

(2 citation statements)

References 29 publications

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“…Firstly, through molecular dynamics, the single mutant (Q157H) with higher thermal stability was identified and then site-directed mutagenesis was employed to create the specific mutation. Compared with the WT, the half-life time of the mutant Q157H was 6.0-fold and 2.08-fold higher than that of the WT at 25 and 37 • C [64].…”

Section: Semi-rational Enzyme Engineering To Increase the Thermostabi...mentioning

confidence: 84%

A Review of Advanced Molecular Engineering Approaches to Enhance the Thermostability of Enzyme Breakers: From Prospective of Upstream Oil and Gas Industry

Naeem

Khalil

Tariq

et al. 2022

IJMS

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During the fracture stimulation of oil and gas wells, fracturing fluids are used to create fractures and transport the proppant into the fractured reservoirs. The fracturing fluid viscosity is responsible for proppant suspension, the viscosity can be increased through the incorporation of guar polymer and cross-linkers. After the fracturing operation, the fluid viscosity is decreased by breakers for efficient oil and gas recovery. Different types of enzyme breakers have been engineered and employed to reduce the fracturing fluid′s viscosity, but thermal stability remains the major constraint for the use of enzymes. The latest enzyme engineering approaches such as direct evolution and rational design, have great potential to increase the enzyme breakers’ thermostability against high temperatures of reservoirs. In this review article, we have reviewed recently advanced enzyme molecular engineering technologies and how these strategies could be used to enhance the thermostability of enzyme breakers in the upstream oil and gas industry.

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Section: Semi-rational Enzyme Engineering To Increase the Thermostabi...mentioning

confidence: 84%

A Review of Advanced Molecular Engineering Approaches to Enhance the Thermostability of Enzyme Breakers: From Prospective of Upstream Oil and Gas Industry

Naeem

Khalil

Tariq

et al. 2022

IJMS

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“…PrChSaseAC showed high activity to CS-A and HA, respectively. Molecular docking showed that more hydrogens were formed in the active pocket among ChSaseAC-CS-C, which limits the extension of the substrate in the active center, resulting in lower enzyme activity ( Li et al, 2019 , Li et al, 2020 ; Su et al, 2022 ), while HA lacks the sulfate moieties of galactosamine, resulting in the decreased ability on the recognition of substrate for enzyme, which makes the enzyme activity slightly lower than using CS-A as the substrate. Asn125, Arg288, Arg292, and Asn374 are involved in the binding of the substrate.…”

Section: Discussionmentioning

confidence: 99%

A Novel Chondroitin AC Lyase With Broad Substrate Specificity From Pedobacter rhizosphaerae: Cloning, Expression, and Characterization

Zhou

Guo

Wei

et al. 2021

Front. Bioeng. Biotechnol.

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Chondroitin AC lyase (ChSaseAC) is one of the essential polysaccharides lyases in low molecular chondroitin sulfate production. In this work, a novel PrChSaseAC from Pedobacter rhizosphaerae was successfully cloned, expressed in Escherichia coli. After optimizing the induction, the recombinant PrChSaseAC could be expressed efficiently at 0.1 mM IPTG, 25°C, and 12 h induction. Then, it was purified with Ni-NTA affinity chromatography. The characterization of the purified PrChSaseAC showed that it had high specific activity and good storage stability, which would favor the production of low molecular weight chondroitin sulfate. It also displayed activity toward chondroitin sulfate C and hyaluronic acid. PrChSaseAC had the highest activity at pH 7.5, 37°C, 10 mM Ca2+, and 5 mg/ml of chondroitin sulfate A. Molecular docking of substrate and enzyme showed the interactions between the enzyme and substrate; it revealed that the enzyme showed high activity to CS-A and hyaluronic acid, but lower activity to CS-C attributed to the structure of the binding pocket. The high stability and specific activity of the enzyme will benefit the industrial production or clinical treatment.

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Engineering of Substrate-Binding Domain to Improve Catalytic Activity of Chondroitin B Lyase with Semi-Rational Design

Tian,

Xu,

et al. 2024

CIMB

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Dermatan sulfate and chondroitin sulfate are dietary supplements that can be utilized as prophylactics against thrombus formation. Low-molecular-weight dermatan sulfate (LMWDS) is particularly advantageous due to its high absorbability. The enzymatic synthesis of low-molecular-weight dermatan sulfates (LMWDSs) using chondroitin B lyase is a sustainable and environmentally friendly approach to manufacturing. However, the industrial application of chondroitin B lyases is severely hampered by their low catalytic activity. To improve the activity, a semi-rational design strategy of engineering the substrate-binding domain of chondroitin B lyase was performed based on the structure. The binding domain was subjected to screening of critical residues for modification using multiple sequence alignments and molecular docking. A total of thirteen single-point mutants were constructed and analyzed to assess their catalytic characteristics. Out of these, S90T, N103C, H134Y, and R159K exhibited noteworthy enhancements in activity. This study also examined combinatorial mutagenesis and found that the mutant H134Y/R159K exhibited a substantially enhanced catalytic activity of 1266.74 U/mg, which was 3.21-fold that of the wild-type one. Molecular docking revealed that the enhanced activity of the mutant could be attributed to the formation of new hydrogen bonds and hydrophobic interactions with the substrate as well as neighbor residues. The highly active mutant would benefit the utilization of chondroitin B lyase in pharmaceuticals and functional foods.

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A single site mutation significantly improves the thermostability and activity of heparinase I fromBacteroides eggerthii

Cited by 3 publications

References 29 publications

A Review of Advanced Molecular Engineering Approaches to Enhance the Thermostability of Enzyme Breakers: From Prospective of Upstream Oil and Gas Industry

A Review of Advanced Molecular Engineering Approaches to Enhance the Thermostability of Enzyme Breakers: From Prospective of Upstream Oil and Gas Industry

A Novel Chondroitin AC Lyase With Broad Substrate Specificity From Pedobacter rhizosphaerae: Cloning, Expression, and Characterization

Engineering of Substrate-Binding Domain to Improve Catalytic Activity of Chondroitin B Lyase with Semi-Rational Design

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