Structure-based protein engineering of bacterial β-xylosidase to increase the production yield of xylobiose from xylose

Hong, Seokho; Kyung, Myungok; Jo, Inseong; Kim, Yong‐Ro; Ha, Nam‐Chul

doi:10.1016/j.bbrc.2018.05.051

Cited by 23 publications

(22 citation statements)

References 21 publications

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“…According to the previous studies of XOs, xylobiose (X2), and xylopentaose (X5) were found to be good nutraceuticals, and exhibited better potential prebiotic properties. X2 shows the highest prebiotic ability among XOs and is better than that of other non‐digestible oligosaccharides for the proliferation of Bifidobacteria in the human gut . Similar to X2, X5 is also important for prebiotic applications; moreover, some researchers have demonstrated that X5 still exhibited a higher anti‐cancer activity .…”

Section: Introductionmentioning

confidence: 99%

Characterization of a novel xylanase from Aspergillus flavus with the unique properties in production of xylooligosaccharides

et al. 2019

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A novel xylanase from the filamentous fungus Aspergillus flavus was purified and characterized as the β‐1, 4‐endoxylanase (designed as AfXynB) with a molecular mass (32.2 kDa), which is different from all of the previously reported xylanases from the same strain. AfXynB was optimally active at pH 7.5 and 55 °C, respectively. It was stable up to 50 °C within range of pH 4.0–9.5, and displayed an excellent tolerance to various cations, reagents, and proteases. AfXynB showed specific activity toward beechwood xylan but no detected activity toward CMC and pNP‐β‐D‐xylopyranoside. The xylanase is a typical endo‐xylanase; it could hydrolyze beechwood xylan to only yield xylobiose (X2) and xylopentaose (X5). Actually, this may be the first report for the endo‐xylanases that displayed such a unique hydrolytic property. These findings in the present study have great implications for its future applications of the novel xylanase.

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Section: Introductionmentioning

confidence: 99%

Characterization of a novel xylanase from Aspergillus flavus with the unique properties in production of xylooligosaccharides

et al. 2019

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“…Lastly, the ab initio SAXS generated envelope shows a good shape agreement with the crystallographic dimer (Figure c). Frequently, tetramers are predicted as the most stable oligomeric form for GH43 subfamily 11 members and have already been observed in solution in equilibrium with a population of dimers (Brunzelle, Jordan, McCaslin, Olczak, & Wawrzak, ; Hong, Kyung, Jo, Kim, & Ha, ). Despite already structurally characterized enzymes from subfamily 12 being predicted by PDBePISA (Krissinel & Henrick, ) as monomers (Hemsworth et al, ) or dimer (Rohman, van Oosterwijk, Puspaningsih, & Dijkstra, ), there was no experimental evidence of their oligomeric state so far.…”

Section: Resultsmentioning

confidence: 99%

“…Structures chosen for comparisons are from distinct organisms and are organized in decreasing order of sequence identity with BlXynB. (PDB ID 1YRZ: Bacillus halodurans [Federov, Federov, & Almo, ]; 2EXH: Geobacillus stearothemophilus [Brüx et al, ]; 3C2U: Selenomonas ruminantium [Brunzelle et al, ]; 5JOW: Bacteriodes ovatus BoGH43A [Hemsworth et al, ]; 5Z5D: Geobacillus thermoleovorans [Rohman et al, ]; 5ZQJ: Bacillus pumilus [Hong et al, ]). BlXynB: Bacillus licheniformis exo‐β‐1,4‐xylanase; GH43: glycoside hydrolase family 43; PDB: Protein Data Bank; UV: ultraviolet [Color figure can be viewed at wileyonlinelibrary.com]…”

Section: Resultsmentioning

confidence: 99%

“…(b) Structure‐based sequence alignment of BlXynB with the most similar structure deposited on PDB (1YRZ from Bacillus halodurans predicted as β‐xylosidase (Federov et al, ) and with structurally and functionally characterized members of subfamilies 11 and 12 (identified by PDB codes). Structures chosen for comparison are from distinct organisms (PDB ID 2EXH: Geobacillus stearothemophilus [Brüx et al, ]; 3C2U: Selenomonas ruminantium [Brunzelle et al, ]; 5JOW: Bacteriodes ovatus BoGH43A [Hemsworth et al, ]; 5Z5D: Geobacillus thermoleovorans [Rohman et al, ]; 5ZQJ: Bacillus pumilus [Hong et al, ]). For Bacteriodes ovatus , the most active GH43 β‐xylosidase (Hemsworth et al, ) from the subfamily 12 was chosen.…”

Section: Resultsmentioning

confidence: 99%

“…Another important point highlighted in this study regards the BlXynB quaternary structure and whether this could affect protein activity. The oligomeric states of only two members of subfamily 11 have been validated in solution (Brunzelle et al, ; Hong et al, ). In both cases, it was observed a mixture of tetramers and dimers, being the tetramers the most stable oligomeric form, according to PDBePISA (Krissinel & Henrick, ).…”

Section: Discussionmentioning

confidence: 99%

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Structure‐guided design combined with evolutionary diversity led to the discovery of the xylose‐releasing exo‐xylanase activity in the glycoside hydrolase family 43

Zanphorlin

Morais

Diogo

et al. 2019

Biotech & Bioengineering

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Rational design is an important tool for sculpting functional and stability properties of proteins and its potential can be much magnified when combined with in vitro and natural evolutionary diversity. Herein, we report the structure‐guided design of a xylose‐releasing exo‐β‐1,4‐xylanase from an inactive member of glycoside hydrolase family 43 (GH43). Structural analysis revealed a nonconserved substitution (Lys247) that results in the disruption of the hydrogen bond network that supports catalysis. The mutation of this residue to a conserved serine restored the catalytic activity and crystal structure elucidation of the mutant confirmed the recovery of the proper orientation of the catalytically relevant histidine. Interestingly, the tailored enzyme can cleave both xylooligosaccharides and xylan, releasing xylose as the main product, being the first xylose‐releasing exo‐β‐1,4‐xylanase reported in the GH43 family. This enzyme presents a unique active‐site topology when compared with closely related β‐xylosidases, which is the absence of a hydrophobic barrier at the positive‐subsite region, allowing the accommodation of long substrates. Therefore, the combination of rational design for catalytic activation along with naturally occurring differences in the substrate binding interface led to the discovery of a novel activity within the GH43 family. In addition, these results demonstrate the importance of solvation of the β‐propeller hollow for GH43 catalytic function and expand our mechanistic understanding about the diverse modes of action of GH43 members, a key and polyspecific carbohydrate‐active enzyme family abundant in most plant cell‐wall‐degrading microorganisms.

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Enzyme engineering: Reshaping the biocatalytic functions

Ali

Ishqi

Husain

2020

Biotech & Bioengineering

124

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Enzyme engineering is a powerful tool to fine‐tune the enzymes. It is a technique by which the stability, activity, and specificity of the enzymes can be altered. The characteristic properties of an enzyme can be amended by immobilization and protein engineering. Among them, protein engineering is the most promising, as in addition to amending the stability and activity, it is the only way to modulate the specificity and stereoselectivity of enzymes. The current review sheds light on protein engineering and the approaches applied for it on the basis of the degree of knowledge of structure and function of enzymes. Enzymes, which have been engineered are also discussed in detail and categorized on the basis of their respective applications. This will give a better insight into the revolutionary changes brought by protein engineering of enzymes in various industrial and environmental processes.

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Structure-based protein engineering of bacterial β-xylosidase to increase the production yield of xylobiose from xylose

Cited by 23 publications

References 21 publications

Characterization of a novel xylanase from Aspergillus flavus with the unique properties in production of xylooligosaccharides

Characterization of a novel xylanase from Aspergillus flavus with the unique properties in production of xylooligosaccharides

Structure‐guided design combined with evolutionary diversity led to the discovery of the xylose‐releasing exo‐xylanase activity in the glycoside hydrolase family 43

Enzyme engineering: Reshaping the biocatalytic functions

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