Functional analysis of chimeric TrCel6A enzymes with different carbohydrate binding modules

Christensen, Stefan Jarl; Badino, Silke Flindt; Cavaleiro, Ana Mafalda; Borch, Kim; Westh, Peter

doi:10.1093/protein/gzaa003

Cited by 8 publications

(6 citation statements)

References 66 publications

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“…A comparative biochemical study of GH6 CBHs from different fungi has shown variation in substrate interactions, adsorption capacity, and turnover number [ 61 ]. Variation in CBD sequence has further been shown to influence the adsorption and kinetics of GH6 CBH [ 62 ]. Taken together, the results of this and previous studies indicate the potential role of cellulase sequence and domain organization in host preferences of pathogenic fungi [ 39 , 62 , 63 , 64 ].…”

Section: Resultsmentioning

confidence: 99%

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Comparative Analysis of Herbaceous and Woody Cell Wall Digestibility by Pathogenic Fungi

et al. 2021

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Fungal pathogens have evolved combinations of plant cell-wall-degrading enzymes (PCWDEs) to deconstruct host plant cell walls (PCWs). An understanding of this process is hoped to create a basis for improving plant biomass conversion efficiency into sustainable biofuels and bioproducts. Here, an approach integrating enzyme activity assay, biomass pretreatment, field emission scanning electron microscopy (FESEM), and genomic analysis of PCWDEs were applied to examine digestibility or degradability of selected woody and herbaceous biomass by pathogenic fungi. Preferred hydrolysis of apple tree branch, rapeseed straw, or wheat straw were observed by the apple-tree-specific pathogen Valsa mali, the rapeseed pathogen Sclerotinia sclerotiorum, and the wheat pathogen Rhizoctonia cerealis, respectively. Delignification by peracetic acid (PAA) pretreatment increased PCW digestibility, and the increase was generally more profound with non-host than host PCW substrates. Hemicellulase pretreatment slightly reduced or had no effect on hemicellulose content in the PCW substrates tested; however, the pretreatment significantly changed hydrolytic preferences of the selected pathogens, indicating a role of hemicellulose branching in PCW digestibility. Cellulose organization appears to also impact digestibility of host PCWs, as reflected by differences in cellulose microfibril organization in woody and herbaceous PCWs and variation in cellulose-binding domain organization in cellulases of pathogenic fungi, which is known to influence enzyme access to cellulose. Taken together, this study highlighted the importance of chemical structure of both hemicelluloses and cellulose in host PCW digestibility by fungal pathogens.

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Section: Resultsmentioning

confidence: 99%

“…Variation in CBD sequence has further been shown to influence the adsorption and kinetics of GH6 CBH [ 62 ]. Taken together, the results of this and previous studies indicate the potential role of cellulase sequence and domain organization in host preferences of pathogenic fungi [ 39 , 62 , 63 , 64 ].…”

Section: Resultsmentioning

confidence: 99%

Comparative Analysis of Herbaceous and Woody Cell Wall Digestibility by Pathogenic Fungi

et al. 2021

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“…A chimeric enzyme is commonly generated by fusing the GH catalytic domain obtained from one species with another protein domain such as a CBM from another species. In several studies, chimeric enzymes have been shown to improve the catalytic efficiency, thermostability, as well as substrate specificity (Oliveira et al, 2015;Saadat, 2017;Christensen et al, 2020;Gilmore et al, 2020).…”

Section: Designing New Chimeric Enzymes Inspired By the Domain Architmentioning

confidence: 99%

Carbohydrate Binding Modules: Diversity of Domain Architecture in Amylases and Cellulases From Filamentous Microorganisms

Sidar

Albuquerque²,

Voshol³

et al. 2020

Front. Bioeng. Biotechnol.

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Enzymatic degradation of abundant renewable polysaccharides such as cellulose and starch is a field that has the attention of both the industrial and scientific community. Most of the polysaccharide degrading enzymes are classified into several glycoside hydrolase families. They are often organized in a modular manner which includes a catalytic domain connected to one or more carbohydrate-binding modules. The carbohydrate-binding modules (CBM) have been shown to increase the proximity of the enzyme to its substrate, especially for insoluble substrates. Therefore, these modules are considered to enhance enzymatic hydrolysis. These properties have played an important role in many biotechnological applications with the aim to improve the efficiency of polysaccharide degradation. The domain organization of glycoside hydrolases (GHs) equipped with one or more CBM does vary within organisms. This review comprehensively highlights the presence of CBM as ancillary modules and explores the diversity of GHs carrying one or more of these modules that actively act either on cellulose or starch. Special emphasis is given to the cellulase and amylase distribution within the filamentous microorganisms from the genera of Streptomyces and Aspergillus that are well known to have a great capacity for secreting a wide range of these polysaccharide degrading enzyme. The potential of the CBM and other ancillary domains for the design of improved polysaccharide decomposing enzymes is discussed.

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“…S2 and the corresponding parameters are summarized in Table 1 . B max / K d was applied to quantify the affinity of the protein to cellulose (Arola and Linder 2016 ; Christensen et al 2020 ). The B max / K d of Pp-ArCel5 and Pp-ArCel5-NG towards filter paper (CrI ≈ 84%) was significantly higher than that towards Avicel (CrI ≈ 77%), due to difference crystallinity of substrates (Hall et al 2010 ).…”

Section: Resultsmentioning

confidence: 99%

A novel accessory protein ArCel5 from cellulose-gelatinizing fungus Arthrobotrys sp. CX1

Yuan

Chen

Wang

et al. 2022

Bioresour. Bioprocess.

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Improved understanding of cellulose swelling mechanism is beneficial for increasing the hydrolysis efficiency of cellulosic substrates. Here, we report a family 5 glycoside hydrolase ArCel5 isolated from the cellulose-gelatinizing fungus Arthrobotrys sp. CX1. ArCel5 exhibited low specific hydrolysis activity and high cellulose swelling capability, which suggested that this protein might function as an accessory protein. Homology modeling glycosylation detection revealed that ArCel5 is a multi-domain protein including a family 1 carbohydrate-binding module, a glycosylation linker, and a catalytic domain. The adsorption capacity, structural changes and hydrature index of filter paper treated by different ArCel5 mutants demonstrated that CBM1 and linker played an essential role in recognizing, binding and decrystallizing cellulosic substrates, which further encouraged the synergistic action between ArCel5 and cellulases. Notably, glycosylation modification further strengthened the function of the linker region. Overall, our study provides insight into the cellulose decrystallization mechanism by a novel accessory protein ArCel5 that will benefit future applications. Graphical Abstract

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Functional analysis of chimeric TrCel6A enzymes with different carbohydrate binding modules

Cited by 8 publications

References 66 publications

Comparative Analysis of Herbaceous and Woody Cell Wall Digestibility by Pathogenic Fungi

Comparative Analysis of Herbaceous and Woody Cell Wall Digestibility by Pathogenic Fungi

Carbohydrate Binding Modules: Diversity of Domain Architecture in Amylases and Cellulases From Filamentous Microorganisms

A novel accessory protein ArCel5 from cellulose-gelatinizing fungus Arthrobotrys sp. CX1

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