Improving the specific activity of β-mannanase from Aspergillus niger BK01 by structure-based rational design

Huang, Jian‐Wen; Chen, Chun‐Chi; Huang, Chun‐Hsiang; Huang, Ting-Yung; Wu, Tzu-Hui; Cheng, Ya‐Shan; Ko, Tzu‐Ping; Lin, Cheng-Yen; Liu, Je-Ruei; Guo, Rey‐Ting

doi:10.1016/j.bbapap.2014.01.011

Cited by 38 publications

(31 citation statements)

References 40 publications

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“…After cultivation for approximately 168 h, the enzyme activity reached 17 500 U ml −1 culture. This level was apparently higher than the strains of Aspergillus flavus generated by the UV mutagenesis method (Olaniyi et al ), and also higher than those simply cloning mannanase genes from A. niger and Rhizomucor miehei and heterologously expressed in P. pastoris (Do et al ; Li et al ), and even higher than the structure‐based rational design mannanase from A. niger (Huang et al ).…”

Section: Discussionmentioning

confidence: 89%

“…from natural environments, UV-mediated mutagenesis and optimizing cultivation conditions have been used to acquire strains that produce high levels of mannanase (Olaniyi et al 2014;Elsharouny et al 2015;Shimizu et al 2015;Ahirwar et al 2016). Currently, genetic engineering techniques are more popularly used by researchers for microbe breeding (Mao and Zhang 2006;Huang et al 2014;Li et al 2018), and improving the gene dosage in the host genome has been proven to be an efficient method to obtain a high expression level of the target gene (Men endez et al 2000;Shu et al 2016).…”

Section: Introductionmentioning

confidence: 99%

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Manno‐oligosaccharide preparation by the hydrolysis of konjac flour with a thermostable endo‐mannanase from Talaromyces cellulolyticus

Yang

Chen

Zhou

et al. 2019

J of Applied Microbiology

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Aims A thermostable endo‐mannanase from the fungus Talaromyces cellulolyticus was identified to facilitate manno‐oligosaccharide preparation from Konjac (Amorphophallus konjac) flour. Methods and Results A putative endo‐1,4‐β‐mannanase from the T. cellulolyticus was obtained and efficiently expressed by improving its gene dosage in the genome of the host. After cultivation in a bench‐top bioreactor for about 120 h, the protein content and enzyme activity of mannanase increased to 3·4 g l−1 and 17 500 U ml−1 respectively. Enzymatic characterization showed that this enzyme has an optimal temperature of 80°C, optimal pH of 5·0. Under the optimized hydrolysis conditions of pH 5·0, 70°C, and an enzyme concentration of 200 U l−1 solution, this enzyme could efficiently hydrolyse 0·5% konjac flour into manno‐oligosaccharides (MOSs) with the degree of polymerization range from 3 to 7. The possible mechanism by which the enzyme produced MOSs was also discussed. Conclusion Talaromyces cellulolyticus endo‐mannanase is thermostable and has a broad pH range adaptability. Method of improving the dosage of mannanase gene in the genome could realized its high‐level impression. This enzyme could efficiently hydrolyse konjac flour into manno‐oligosaccharide products. Significance and Impact of the Study This study has enriched endo‐mannanase resources, facilitated its bulk production and provided a strong reference for its application in manno‐oligosaccharide preparation from the natural glucomannan of konjac flour.

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Section: Discussionmentioning

confidence: 89%

Section: Introductionmentioning

confidence: 99%

Manno‐oligosaccharide preparation by the hydrolysis of konjac flour with a thermostable endo‐mannanase from Talaromyces cellulolyticus

Yang

Chen

Zhou

et al. 2019

J of Applied Microbiology

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“…Trp-125 is located at the reducing end of the active site to provide a stacking force to the +2 sugar. We intended to design mutants W125F and W125A to facilitate the release of cleaved products, the strategy successfully applied to other glycoside hydrolases Huang et al 2014). From the mutant results of W125A, which showed lower activity than the wild-type enzyme, and W125F, whose activity was higher, the stacking force might be required in this position.…”

Section: Discussionmentioning

confidence: 99%

“…In comparison, rational design is more accurate and efficient, by which the desirable features can be introduced to target sites based on protein structural analyses (Labrou 2010;Romero and Arnold 2009). In our previous studies, we have conducted structure-based rational design and successfully improved catalytic efficacy and thermostability of several enzymes Huang et al 2012Huang et al , 2014Wu et al 2014).…”

Section: Introductionmentioning

confidence: 98%

Improving the catalytic performance of a GH11 xylanase by rational protein engineering

Cheng

Chen

Huang

et al. 2015

Appl Microbiol Biotechnol

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XynCDBFV from Neocallimastix patriciarum is among the most effective xylanases and holds great potentials in a wide variety of industrial applications. In the present study, several active site residues were modified referring to the instrumental information of the complex structure of XynCDBFV and xylooligosaccharides. Among the 12 single active site mutants, W125F and F163W show increased activity comparing to the wild-type protein. The double mutant W125F/F163W was then generated which displayed nearly 20 % increase in the enzyme activity. Although W125F/F163W showed 5 °C reduction in the optimal temperature, it still preserves similar thermostability and is more active than the wild-type enzyme at temperatures lower than 60 °C. These properties make the double mutant a suitable candidate for commercial applications that involve lower operating temperatures. Furthermore, we investigated the effect of N-glycosylation on the thermostability of XynCDBFV when expressed in the yeast strain Pichia pastoris for industrial use. Two potential glycosylation sites (Asn-37 and Asn-88) were examined, and their roles in enzyme performance were validated. We found that the N-glycosylations of XynCDBFV are related to both catalytic activity and heat stability, with Asn-37 motif playing a dominant role. Collectively, the enzymatic properties of XynCDBFV were improved by molecular engineering, and the influences of N-glycosylations on the enzyme have been clearly elucidated herein.

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“…This glycosylation may contribute to MANF3 stability. 16) At present, the N-glycosylation pattern of the recombinant MANF3 cannot be directly compared to that of the native enzyme, since the glycosylation of the native enzyme has not been tested.…”

Section: Discussionmentioning

confidence: 99%

Production of high activity Aspergillus niger BCC4525 β-mannanase in Pichia pastoris and its application for mannooligosaccharides production from biomass hydrolysis

Harnpicharnchai

Pinngoen

Teanngam

et al. 2016

Bioscience, Biotechnology, and Biochemistry

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A cDNA encoding β-mannanase was cloned from Aspergillus niger BCC4525 and expressed in Pichia pastoris KM71. The secreted enzyme hydrolyzed locust bean gum substrate with very high activity (1625 U/mL) and a relatively high k/K (461 mg s mL). The enzyme is thermophilic and thermostable with an optimal temperature of 70 °C and 40% retention of endo-β-1,4-mannanase activity after preincubation at 70 °C. In addition, the enzyme exhibited broad pH stability with an optimal pH of 5.5. The recombinant enzyme hydrolyzes low-cost biomass, including palm kernel meal (PKM) and copra meal, to produce mannooligosaccharides, which is used as prebiotics to promote the growth of beneficial microflora in animals. An in vitro digestibility test simulating the gastrointestinal tract system of broilers suggested that the recombinant β-mannanase could effectively liberate reducing sugars from PKM-containing diet. These characteristics render this enzyme suitable for utilization as a feed additive to improve animal performance.

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Improving the specific activity of β-mannanase from Aspergillus niger BK01 by structure-based rational design

Cited by 38 publications

References 40 publications

Manno‐oligosaccharide preparation by the hydrolysis of konjac flour with a thermostable endo‐mannanase from Talaromyces cellulolyticus

Manno‐oligosaccharide preparation by the hydrolysis of konjac flour with a thermostable endo‐mannanase from Talaromyces cellulolyticus

Improving the catalytic performance of a GH11 xylanase by rational protein engineering

Production of high activity Aspergillus niger BCC4525 β-mannanase in Pichia pastoris and its application for mannooligosaccharides production from biomass hydrolysis

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