Improvement in activity of cellulase Cel12A of Thermotoga neapolitana by error prone PCR

Basit, Abdul; Tajwar, Razia; Sadaf, Saima; Zhang, Yang; Akhtar, Muhammad

doi:10.1016/j.jbiotec.2019.09.011

Cited by 21 publications

(5 citation statements)

References 28 publications

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“…Since, there was no PTM site predicted in tACE2, therefore, E. coli would be an ideal host for its large scale expression. E. coli is the easiest, quickest, and cheapest expression host with a fully known genome, most widely used for hetrologous expression of recombinant protein (Basit et al, 2019). Since, ACE2 is eukaryotic protein; therefore, its expression in its native form in E. coli will be uncertain, as most of the eukaryotic protein showed insoluble expression in E. coli, which need to be refolded invitro , which is costly and time consuming.…”

Section: Tace2 Predicted Soluble Expression In E Colimentioning

confidence: 99%

Truncated human angiotensin converting enzyme 2; a potential inhibitor of SARS-CoV-2 spike glycoprotein and potent COVID-19 therapeutic agent

Basit

Ali

Rehman

2020

Journal of Biomolecular Structure and Dynamics

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The current pandemic of Covid-19 caused by SARS-CoV-2 is continued to spread globally and no potential drug or vaccine against it is available. Spike (S) glycoprotein is the structural protein of SARS-CoV-2 located on the envelope surface, involve in interaction with angiotensin converting enzyme 2 (ACE2), a cell surface receptor, followed by entry into the host cell. Thereby, blocking the S glycoprotein through potential inhibitor may interfere its interaction with ACE2 and impede its entry into the host cell. Here, we present a truncated version of human ACE2 (tACE2), comprising the N terminus region of the intact ACE2 from amino acid position 21-119, involved in binding with receptor binding domain (RBD) of SARS-CoV-2. We analyzed the in-silico potential of tACE2 to compete with intact ACE2 for binding with RBD. The protein-protein docking and molecular dynamic simulation showed that tACE2 has higher binding affinity for RBD and form more stabilized complex with RBD than the intact ACE2. Furthermore, prediction of tACE2 soluble expression in E. coli makes it a suitable candidate to be targeted for Covid-19 therapeutics. This is the first MD simulation based findings to provide a high affinity protein inhibitor for SARS-CoV-2 S glycoprotein, an important target for drug designing against this unprecedented challenge.

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Section: Tace2 Predicted Soluble Expression In E Colimentioning

confidence: 99%

Truncated human angiotensin converting enzyme 2; a potential inhibitor of SARS-CoV-2 spike glycoprotein and potent COVID-19 therapeutic agent

Basit

Ali

Rehman

2020

Journal of Biomolecular Structure and Dynamics

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“…In 1978, Michael Smith first proposed site-specific mutagenesis, which opened the door to protein modification and design . Since then, a series of classical mutagenesis methods have been developed and applied, mainly including saturation mutagenesis (SM), error-prone PCR ( ep PCR), and DNA shuffling …”

Section: Tunnel Engineering Of Lipasementioning

confidence: 99%

“…90 In 1978, Michael Smith first proposed sitespecific mutagenesis, which opened the door to protein modification and design. 91 Since then, a series of classical mutagenesis methods have been developed and applied, mainly including saturation mutagenesis (SM), 92 error-prone PCR (epPCR), 93 and DNA shuffling. 94 Based on a phenol red high-throughput screening method, Liu et al used epPCR to improve the catalytic activity of D-carbamoylase for D-amino acids and revealed that the variant R211G exhibited significantly higher activity compared to the wild-type (WT).…”

Section: Directed Evolution Of Tunnelmentioning

confidence: 99%

Computer-Aided Tunnel Engineering: A Promising Strategy for Improving Lipase Applications in Esterification Reactions

Chong,

Qi,

et al. 2023

ACS Catal.

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Esterification is one of the most important reactions in organic synthesis and industrial applications. The applications of lipases in esterification are increasingly widespread and have high cumulative values. Mostly, in the catalytic cycle of lipases, the tunnel connecting the active site to the surface of a protein plays a significant role in ligand transport. As a unique conformational structure of a protein, the tunnel could regulate the catalytic performance of lipases via ligand transportation. Therefore, the tunnel of lipase can be designed accordingly to speed up the transportation of the ligand, also highlighting the importance of tunnel engineering in the industrial application of enzymatic esterification reactions. This article reviewed the characterizations of the tunnel structure and focused on the strategies of tunnel engineering, such as rational design and directed evolution. Additionally, tunnel engineering using computer-aided design techniques (e.g., CAVER, Molecular Dynamics simulations) to improve the catalytic functions of lipases, such as activity, substrate specificity, and stability, was also discussed in depth. Finally, this review provides future perspectives about tunnel-engineered lipases as biocatalysts in esterification reactions.

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“…Similarly, the specific activity of Bacillus stearothermophilus xylanase A (BaxA) was increased by 3.5-fold (9.38 Umg −1 ) using directed evolution ( Xu et al, 2016 ). The cellulase activity of a recombinant enzyme from Thermotoga neapolitana was enhanced using Random mutagenesis, the best mutants obtained when compared to the wild type shows 2.7-, 5- and 4.8- fold increase in activity against CMC, RAC and Avicel respectively ( Basit et al, 2019 ). The use of mutations is an accepted method to improve recombinant proteins at a genetic level, although screening hundreds or thousands of mutants is tedious ( Stephens et al, 2014 ).…”

Section: Strategies To Enhance Thermostability and Specific Activity Of Cellulase And Xylanase Of Thermophilesmentioning

confidence: 99%

Thermostable Cellulases / Xylanases From Thermophilic and Hyperthermophilic Microorganisms: Current Perspective

Ajeje¹,

Hu²,

Song³

et al. 2021

Front. Bioeng. Biotechnol.

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The bioconversion of lignocellulose into monosaccharides is critical for ensuring the continual manufacturing of biofuels and value-added bioproducts. Enzymatic degradation, which has a high yield, low energy consumption, and enhanced selectivity, could be the most efficient and environmentally friendly technique for converting complex lignocellulose polymers to fermentable monosaccharides, and it is expected to make cellulases and xylanases the most demanded industrial enzymes. The widespread nature of thermophilic microorganisms allows them to proliferate on a variety of substrates and release substantial quantities of cellulases and xylanases, which makes them a great source of thermostable enzymes. The most significant breakthrough of lignocellulolytic enzymes lies in lignocellulose-deconstruction by enzymatic depolymerization of holocellulose into simple monosaccharides. However, commercially valuable thermostable cellulases and xylanases are challenging to produce in high enough quantities. Thus, the present review aims at giving an overview of the most recent thermostable cellulases and xylanases isolated from thermophilic and hyperthermophilic microbes. The emphasis is on recent advancements in manufacturing these enzymes in other mesophilic host and enhancement of catalytic activity as well as thermostability of thermophilic cellulases and xylanases, using genetic engineering as a promising and efficient technology for its economic production. Additionally, the biotechnological applications of thermostable cellulases and xylanases of thermophiles were also discussed.

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Improvement in activity of cellulase Cel12A of Thermotoga neapolitana by error prone PCR

Cited by 21 publications

References 28 publications

Truncated human angiotensin converting enzyme 2; a potential inhibitor of SARS-CoV-2 spike glycoprotein and potent COVID-19 therapeutic agent

Truncated human angiotensin converting enzyme 2; a potential inhibitor of SARS-CoV-2 spike glycoprotein and potent COVID-19 therapeutic agent

Computer-Aided Tunnel Engineering: A Promising Strategy for Improving Lipase Applications in Esterification Reactions

Thermostable Cellulases / Xylanases From Thermophilic and Hyperthermophilic Microorganisms: Current Perspective

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