Additional Carbohydrate-Binding Modules Enhance the Insoluble Substrate-Hydrolytic Activity of β-1,3-Glucanase from AlkaliphilicNocardiopsissp. F96

2009 ◽  
Vol 73 (5) ◽  
pp. 1078-1082 ◽  
Author(s):  
Naoya KOIZUMI ◽  
Sumiko MASUDA ◽  
Kiyoe MAEDA ◽  
Yuya ISODA ◽  
Rie YATSUNAMI ◽  
...  
Keyword(s):  
Hydrolytic Activity ◽  
Carbohydrate Binding ◽  
Insoluble Substrate ◽  
Carbohydrate Binding Modules

scholarly journals A trimodular bacterial enzyme combining hydrolytic activity with oxidative glycosidic bond cleavage efficiently degrades chitin

2020 ◽  
Vol 295 (27) ◽  
pp. 9134-9146 ◽  
Author(s):  
Sophanit Mekasha ◽  
Tina Rise Tuveng ◽  
Fatemeh Askarian ◽  
Swati Choudhary ◽  
Claudia Schmidt-Dannert ◽  
...  
Keyword(s):  
Catalytic Domain ◽  
Bond Cleavage ◽  
Hydrolytic Enzymes ◽  
Hydrolytic Activity ◽  
Full Length ◽  
Spatial Proximity ◽  
Carbohydrate Binding ◽  
Lytic Polysaccharide Monooxygenase ◽  
Chitin Binding Domain ◽  
Carbohydrate Binding Modules

Findings from recent studies have indicated that enzymes containing more than one catalytic domain may be particularly powerful in the degradation of recalcitrant polysaccharides such as chitin and cellulose. Some known multicatalytic enzymes contain several glycoside hydrolase domains and one or more carbohydrate-binding modules (CBMs). Here, using bioinformatics and biochemical analyses, we identified an enzyme, Jd1381 from the actinobacterium Jonesia denitrificans, that uniquely combines two different polysaccharide-degrading activities. We found that Jd1381 contains an N-terminal family AA10 lytic polysaccharide monooxygenase (LPMO), a family 5 chitin-binding domain (CBM5), and a family 18 chitinase (Chi18) domain. The full-length enzyme, which seems to be the only chitinase produced by J. denitrificans, degraded both α- and β-chitin. Both the chitinase and the LPMO activities of Jd1381 were similar to those of other individual chitinases and LPMOs, and the overall efficiency of chitin degradation by full-length Jd1381 depended on its chitinase and LPMO activities. Of note, the chitin-degrading activity of Jd1381 was comparable with or exceeded the activities of combinations of well-known chitinases and an LPMO from Serratia marcescens. Importantly, comparison of the chitinolytic efficiency of Jd1381 with the efficiencies of combinations of truncated variants—JdLPMO10 and JdCBM5-Chi18 or JdLPMO10-CBM5 and JdChi18—indicated that optimal Jd1381 activity requires close spatial proximity of the LPMO10 and the Chi18 domains. The demonstration of intramolecular synergy between LPMOs and hydrolytic enzymes reported here opens new avenues toward the development of efficient catalysts for biomass conversion.

scholarly journals Functional diversity of three tandem C-terminal carbohydrate-binding modules of a β-mannanase

2021 ◽  
pp. 100638
Author(s):  
Marie Sofie Møller ◽  
Souad El Bouaballati ◽  
Bernard Henrissat ◽  
Birte Svensson
Keyword(s):  
Functional Diversity ◽  
Carbohydrate Binding ◽  
Carbohydrate Binding Modules

scholarly journals Functionalization of Cellulose-Based Hydrogels with Bi-Functional Fusion Proteins Containing Carbohydrate-Binding Modules

Materials ◽  
2021 ◽  
Vol 14 (12) ◽  
pp. 3175
Author(s):  
Mariana Barbosa ◽  
Hélvio Simões ◽  
Duarte Miguel F. Prazeres
Keyword(s):  
Fusion Proteins ◽  
Fluorescent Protein ◽  
Protein A ◽  
Chemical Properties ◽  
Main Idea ◽  
Bioactive Molecules ◽  
Scaffolding Protein ◽  
Carbohydrate Binding ◽  
Biological Interactions ◽  
Carbohydrate Binding Modules

Materials with novel and enhanced functionalities can be obtained by modifying cellulose with a range of biomolecules. This functionalization can deliver tailored cellulose-based materials with enhanced physical and chemical properties and control of biological interactions that match specific applications. One of the foundations for the success of such biomaterials is to efficiently control the capacity to combine relevant biomolecules into cellulose materials in such a way that the desired functionality is attained. In this context, our main goal was to develop bi-functional biomolecular constructs for the precise modification of cellulose hydrogels with bioactive molecules of interest. The main idea was to use biomolecular engineering techniques to generate and purify different recombinant fusions of carbohydrate binding modules (CBMs) with significant biological entities. Specifically, CBM-based fusions were designed to enable the bridging of proteins or oligonucleotides with cellulose hydrogels. The work focused on constructs that combine a family 3 CBM derived from the cellulosomal-scaffolding protein A from Clostridium thermocellum (CBM3) with the following: (i) an N-terminal green fluorescent protein (GFP) domain (GFP-CBM3); (ii) a double Z domain that recognizes IgG antibodies; and (iii) a C-terminal cysteine (CBM3C). The ability of the CBM fusions to bind and/or anchor their counterparts onto the surface of cellulose hydrogels was evaluated with pull-down assays. Capture of GFP-CBM3 by cellulose was first demonstrated qualitatively by fluorescence microscopy. The binding of the fusion proteins, the capture of antibodies (by ZZ-CBM3), and the grafting of an oligonucleotide (to CBM3C) were successfully demonstrated. The bioactive cellulose platform described here enables the precise anchoring of different biomolecules onto cellulose hydrogels and could contribute significatively to the development of advanced medical diagnostic sensors or specialized biomaterials, among others.

scholarly journals Integrated Counts of Carbohydrate-Active Protein Domains as Metabolic Readouts to Distinguish Probiotic Biology and Human Fecal Metagenomes

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Hong-Hsing Liu ◽  
Yu-Chen Lin ◽  
Chen-Shuan Chung ◽  
Kevin Liu ◽  
Ya-Hui Chang ◽  
...  
Keyword(s):  
Markov Models ◽  
Protein Domains ◽  
Degradation Pathway ◽  
The Other ◽  
Carbohydrate Binding ◽  
Gram Stain ◽  
Active Protein ◽  
Metabolic Potential ◽  
Independent Validation ◽  
Carbohydrate Binding Modules

AbstractBowel microbiota is a “metaorgan” of metabolisms on which quantitative readouts must be performed before interventions can be introduced and evaluated. The study of the effects of probiotic Clostridium butyricum MIYAIRI 588 (CBM588) on intestine transplantees indicated an increased percentage of the “other glycan degradation” pathway in 16S-rRNA-inferred metagenomes. To verify the prediction, a scoring system of carbohydrate metabolisms derived from shotgun metagenomes was developed using hidden Markov models. A significant correlation (R = 0.9, p < 0.015) between both modalities was demonstrated. An independent validation revealed a strong complementarity (R = −0.97, p < 0.002) between the scores and the abundance of “glycogen degradation” in bacteria communities. On applying the system to bacteria genomes, CBM588 had only 1 match and ranked higher than the other 8 bacteria evaluated. The gram-stain properties were significantly correlated to the scores (p < 5 × 10−4). The distributions of the scored protein domains indicated that CBM588 had a considerably higher (p < 10−5) proportion of carbohydrate-binding modules than other bacteria, which suggested the superior ability of CBM588 to access carbohydrates as a metabolic driver to the bowel microbiome. These results demonstrated the use of integrated counts of protein domains as a feasible readout for metabolic potential within bacteria genomes and human metagenomes.

scholarly journals Family 6 carbohydrate-binding modules display multiple β1,3-linked glucan-specific binding interfaces

2009 ◽  
Vol 300 (1) ◽  
pp. 48-57 ◽  
Author(s):  
Márcia A.S. Correia ◽  
Virgínia M.R. Pires ◽  
Harry J. Gilbert ◽  
David N. Bolam ◽  
Vânia O. Fernandes ◽  
...  
Keyword(s):  
Specific Binding ◽  
Carbohydrate Binding ◽  
Carbohydrate Binding Modules ◽  
Binding Interfaces

scholarly journals Versatile derivatives of carbohydrate-binding modules for imaging of complex carbohydrates approaching the molecular level of resolution

BioTechniques ◽  
2006 ◽  
Vol 41 (4) ◽  
pp. 435-443 ◽  
Author(s):  
Shi-You Ding ◽  
Qi Xu ◽  
Mursheda K. Ali ◽  
John O. Baker ◽  
Edward A. Bayer ◽  
...  
Keyword(s):  
Molecular Level ◽  
Carbohydrate Binding ◽  
Carbohydrate Binding Modules ◽  
Complex Carbohydrates ◽  
Derivatives Of

scholarly journals Supercharged Cellulases Show Reduced Non-Productive Binding, But Enhanced Activity, on Pretreated Lignocellulosic Biomass

2021 ◽  
Author(s):  
Bhargava Nemmaru ◽  
Jenna Douglass ◽  
John M Yarbrough ◽  
Antonio De Chellis ◽  
Srivatsan Shankar ◽  
...  
Keyword(s):  
Cell Wall ◽  
Corn Stover ◽  
Lignocellulosic Biomass ◽  
Hydrolytic Activity ◽  
Fine Tuning ◽  
Cellulolytic Enzymes ◽  
Crystalline Cellulose ◽  
Cell Wall Components ◽  
Carbohydrate Binding Modules ◽  
Wall Components

Non-productive adsorption of cellulolytic enzymes to various plant cell wall components, such as lignin and cellulose, necessitates high enzyme loadings to achieve efficient conversion of pretreated lignocellulosic biomass to fermentable sugars. Carbohydrate-binding modules (CBMs), appended to various catalytic domains (CDs), promote lignocellulose deconstruction by increasing targeted substrate-bound CD concentration but often at the cost of increased non-productive enzyme binding. Here, we demonstrate how a computational protein design strategy can be applied to a model endocellulase enzyme (Cel5A) from Thermobifida fusca to allow fine-tuning its CBM surface charge, which led to increased hydrolytic activity towards pretreated lignocellulosic biomass (e.g., corn stover) by up to ~330% versus the wild-type Cel5A control. We established that the mechanistic basis for this improvement arises from reduced non-productive binding of supercharged Cel5A mutants to cell wall components such as crystalline cellulose (up to 1.7-fold) and lignin (up to 1.8-fold). Interestingly, supercharged Cel5A mutants that showed improved activity on various forms of pretreated corn stover showed increased reversible binding to lignin (up to 2.2-fold) while showing no change in overall thermal stability remarkably. In general, negative supercharging led to increase hydrolytic activity towards both pretreated lignocellulosic biomass and crystalline cellulose whereas positive supercharging led to a reduction of hydrolytic activity. Overall, selective supercharging of protein surfaces was shown to be an effective strategy for improving hydrolytic performance of cellulolytic enzymes for saccharification of real-world pretreated lignocellulosic biomass substrates. Future work should address the implications of supercharging cellulases from various families on inter-enzyme interactions and synergism.

Novel clostridial cell-surface hemicellulose-binding CBM3 proteins

2021 ◽  
Vol 77 (4) ◽  
Author(s):  
Almog Hershko Rimon ◽  
Oded Livnah ◽  
Inna Rozman Grinberg ◽  
Lizett Ortiz de Ora ◽  
Oren Yaniv ◽  
...  
Keyword(s):  
Calcium Binding ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Carbohydrate Binding ◽  
Unexpected Finding ◽  
Calcium Binding Site ◽  
Three Dimensional Structure ◽  
Loop Region ◽  
Carbohydrate Binding Modules ◽  
Cellulose Binding

A novel member of the family 3 carbohydrate-binding modules (CBM3s) is encoded by a gene (Cthe_0271) in Clostridium thermocellum which is the most highly expressed gene in the bacterium during its growth on several types of biomass substrates. Surprisingly, CtCBM3-0271 binds to at least two different types of xylan, instead of the common binding of CBM3s to cellulosic substrates. CtCBM3-0271 was crystallized and its three-dimensional structure was solved and refined to a resolution of 1.8 Å. In order to learn more about the role of this type of CBM3, a comparative study with its orthologue from Clostridium clariflavum (encoded by the Clocl_1192 gene) was performed, and the three-dimensional structure of CcCBM3-1192 was determined to 1.6 Å resolution. Carbohydrate binding by CcCBM3-1192 was found to be similar to that by CtCBM3-0271; both exhibited binding to xylan rather than to cellulose. Comparative structural analysis of the two CBM3s provided a clear functional correlation of structure and binding, in which the two CBM3s lack the required number of binding residues in their cellulose-binding strips and thus lack cellulose-binding capabilities. This is an enigma, as CtCBM3-0271 was reported to be a highly expressed protein when the bacterium was grown on cellulose. An additional unexpected finding was that CcCBM3-1192 does not contain the calcium ion that was considered to play a structural stabilizing role in the CBM3 family. Despite the lack of calcium, the five residues that form the calcium-binding site are conserved. The absence of calcium results in conformational changes in two loops of the CcCBM3-1192 structure. In this context, superposition of the non-calcium-binding CcCBM3-1192 with CtCBM3-0271 and other calcium-binding CBM3s reveals a much broader two-loop region in the former compared with CtCBM3-0271.

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