Surface charge engineering of β-glucosidase using rational design improves catalytic capacity and ionic liquid tolerance

Mu, Yinghui; Ju, Xin; Fu, Jiaolong; Meng, Fanjin; Lü, Yan; Li, Liangzhi

doi:10.1016/j.molliq.2022.120577

Cited by 11 publications

(7 citation statements)

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“…The negative surface charge improves the ability of halophilic proteins to bind hydrated cations, where they would otherwise disrupt the ordered solvation shell, that help maintain a surface hydration layer, reducing hydrophobicity and preventing denaturation and aggregation 88 . Recent rational design experiments of surface charge engineering of a β-glucosidase have demonstrated these features to have significant effects on stability and catalytic capacity highlighting both the complexity and importance of this characteristic 89 . These features are apparent in SMECel6A which displayed a higher negative surface charge with a greater number of negative and neutral surface residues.…”

Section: Discussionmentioning

confidence: 99%

Functional characterisation of a new halotolerant seawater active glycoside hydrolase family 6 cellobiohydrolase from a salt marsh

Leadbeater,

Bruce

2024

Sci Rep

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Realising a fully circular bioeconomy requires the valorisation of lignocellulosic biomass. Cellulose is the most attractive component of lignocellulose but depolymerisation is inefficient, expensive and resource intensive requiring substantial volumes of potable water. Seawater is an attractive prospective replacement, however seawater tolerant enzymes are required for the development of seawater-based biorefineries. Here, we report a halophilic cellobiohydrolase SMECel6A, identified and isolated from a salt marsh meta-exo-proteome dataset with high sequence divergence to previously characterised cellobiohydrolases. SMECel6A contains a glycoside hydrolase family 6 (GH6) domain and a carbohydrate binding module family 2 (CBM2) domain. Characterisation of recombinant SMECel6A revealed SMECel6A to be active upon crystalline and amorphous cellulose. Mono- and oligosaccharide product profiles revealed cellobiose as the major hydrolysis product confirming SMECel6A as a cellobiohydrolase. We show SMECel6A to be halophilic with optimal activity achieved in 0.5X seawater displaying 80.6 ± 6.93% activity in 1 × seawater. Structural predictions revealed similarity to a characterised halophilic cellobiohydrolase despite sharing only 57% sequence identity. Sequential thermocycling revealed SMECel6A had the ability to partially reversibly denature exclusively in seawater retaining significant activity. Our study confirms that salt marsh ecosystems harbour enzymes with attractive traits with biotechnological potential for implementation in ionic solution based bioprocessing systems.

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Section: Discussionmentioning

confidence: 99%

Functional characterisation of a new halotolerant seawater active glycoside hydrolase family 6 cellobiohydrolase from a salt marsh

Leadbeater,

Bruce

2024

Sci Rep

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“…Maintaining high activity in unconventional phase (e.g., ionic liquid (IL), saline concentrated seawater, or organic solvents) is critical for applications of BGLs in lignocellulosic biocatalysis, saline land improvement, and marine cellulose biomass utilization [ 15 , 117 ]. Therefore, it is important to understand the mechanisms that BGLs can tolerate in the unconventional environment during catalysis and use these mechanisms to guide the discovery of BGLs with higher activity/stability.…”

Section: Engineering Of Bgl Functionalitiesmentioning

confidence: 99%

“…Surface charge engineering is a promising approach from the perspective of IL tolerance. Mutants with increased negative surface charge showed higher catalytic efficiency in IL due to the electrostatic repulsion between IL and the salt-bridge network of BGL [ 15 ]. The salt tolerance of mutants is improved by increasing the acidic amino acids on the protein surface and near the entrance of the active site to hinder the entry of high concentrations of salt ions into the active site [ 13 , 78 ].…”

Section: Engineering Of Bgl Functionalitiesmentioning

confidence: 99%

“…To broaden the applications of BGLs in industry, it would be beneficial to enhance the capability of BGLs to tolerate non-mild conditions such as high temperatures, high concentrations of glucose, extreme pH, and high concentrations of organic solvents, to name a few [ 12 , 13 , 14 , 15 ]. In this paper, we review the recent advances in engineering BGLs with enhanced enzymatic properties.…”

Section: Introductionmentioning

confidence: 99%

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Recent Advances in β-Glucosidase Sequence and Structure Engineering: A Brief Review

et al. 2023

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β-glucosidases (BGLs) play a crucial role in the degradation of lignocellulosic biomass as well as in industrial applications such as pharmaceuticals, foods, and flavors. However, the application of BGLs has been largely hindered by issues such as low enzyme activity, product inhibition, low stability, etc. Many approaches have been developed to engineer BGLs to improve these enzymatic characteristics to facilitate industrial production. In this article, we review the recent advances in BGL engineering in the field, including the efforts from our laboratory. We summarize and discuss the BGL engineering studies according to the targeted functions as well as the specific strategies used for BGL engineering.

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“…To broaden the applications of BGLs in industry, it is beneficial to enhance the capability of BGLs to tolerate non-mild conditions such as high temperatures, high concentrations of glucose, extreme pH, high concentrations of organic solvents, to name a few [12][13][14][15]. In this paper, we review the recent advances in engineering BGLs with enhanced enzymatic properties.…”

Section: Introductionmentioning

confidence: 99%

Recent Advances in β-glucosidase Engineering: A Brief Review

Zhao¹,

Huang²,

Ouyang³

et al. 2023

Preprint

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Surface charge engineering of β-glucosidase using rational design improves catalytic capacity and ionic liquid tolerance

Cited by 11 publications

References 50 publications

Functional characterisation of a new halotolerant seawater active glycoside hydrolase family 6 cellobiohydrolase from a salt marsh

Functional characterisation of a new halotolerant seawater active glycoside hydrolase family 6 cellobiohydrolase from a salt marsh

Recent Advances in β-Glucosidase Sequence and Structure Engineering: A Brief Review

Recent Advances in β-glucosidase Engineering: A Brief Review

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