A novel dimeric exoglucanase (GH5_38): Biochemical and Structural Characterisation towards its Application in Alkyl Cellobioside Synthesis
- Authors: Mafa, Mpho S , Dirr, Heinrich , Malgas, Samkelo , Krause, Rui W M , Pletschke, Brett I
- Date: 2020
- Subjects: To be catalogued
- Language: English
- Type: text , article
- Identifier: http://hdl.handle.net/10962/193976 , vital:45412 , xlink:href="https://doi.org/10.3390/molecules25030746"
- Description: An exoglucanase (Exg-D) from the glycoside hydrolase family 5 subfamily 38 (GH5_38) was heterologously expressed and structurally and biochemically characterised at a molecular level for its application in alkyl glycoside synthesis. The purified Exg-D existed in both dimeric and monomeric forms in solution, which showed highest activity on mixed-linked β-glucan (88.0 and 86.7 U/mg protein, respectively) and lichenin (24.5 and 23.7 U/mg protein, respectively). They displayed a broad optimum pH range from 5.5 to 7 and a temperature optimum from 40 to 60 °C. Kinetic studies demonstrated that Exg-D had a higher affinity towards β-glucan, with a Km of 7.9 mg/mL and a kcat of 117.2 s−1, compared to lichenin which had a Km of 21.5 mg/mL and a kcat of 70.0 s−1. The circular dichroism profile of Exg-D showed that its secondary structure consisted of 11% α-helices, 36% β-strands and 53% coils. Exg-D performed transglycosylation using p-nitrophenyl cellobioside as a glycosyl donor and several primary alcohols as acceptors to produce methyl-, ethyl- and propyl-cellobiosides. These products were identified and quantified via thin-layer chromatography (TLC) and liquid chromatography–mass spectrometry (LC-MS). We concluded that Exg-D is a novel and promising oligomeric glycoside hydrolase for the one-step synthesis of alkyl glycosides with more than one monosaccharide unit.
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- Date Issued: 2020
Evaluating Feruloyl Esterase—Xylanase Synergism for Hydroxycinnamic Acid and Xylo-Oligosaccharide Production from Untreated, Hydrothermally Pre-Treated and Dilute-Acid Pre-Treated Corn Cobs:
- Authors: Mkabayi, Lithalethu , Malgas, Samkelo , Wilhelmi, Brendan S , Pletschke, Brett I
- Date: 2020
- Language: English
- Type: text , article
- Identifier: http://hdl.handle.net/10962/149240 , vital:38818 , https://doi.org/10.3390/agronomy10050688
- Description: Agricultural residues are considered the most promising option as a renewable feedstock for biofuel and high valued-added chemical production due to their availability and low cost. The efficient enzymatic hydrolysis of agricultural residues into value-added products such as sugars and hydroxycinnamic acids is a challenge because of the recalcitrant properties of the native biomass. Development of synergistic enzyme cocktails is required to overcome biomass residue recalcitrance, and achieve high yields of potential value-added products. In this study, the synergistic action of two termite metagenome-derived feruloyl esterases (FAE5 and FAE6), and an endo-xylanase (Xyn11) from Thermomyces lanuginosus, was optimized using 0.5% (w/v) insoluble wheat arabinoxylan (a model substrate) and then applied to 1% (w/v) corn cobs for the efficient production of xylo-oligosaccharides (XOS) and hydroxycinnamic acids.
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- Date Issued: 2020
The Effects of Alkaline Pretreatment on Agricultural Biomasses (Corn Cob and Sweet Sorghum Bagasse) and Their Hydrolysis by a Termite-Derived Enzyme Cocktail:
- Authors: Mafa, Mpho S , Malgas, Samkelo , Bhattacharya, Abhishek , Rashamuse, Konanani , Pletschke, Brett I
- Date: 2020
- Language: English
- Type: text , article
- Identifier: http://hdl.handle.net/10962/160273 , vital:40430 , https://doi.org/10.3390/agronomy10081211
- Description: Sweet sorghum bagasse (SSB) and corncob (CC) have been identified as promising feedstocks for the production of second-generation biofuels and other value-added chemicals. In this study, lime (Ca(OH)2) and NaOH pretreatment efficacy for decreasing recalcitrance from SSB and CC was investigated, and subsequently, the pretreated biomass was subjected to the hydrolytic action of an in-house formulated holocellulolytic enzyme cocktail (HEC-H). Compositional analysis revealed that SSB contained 29.34% lignin, 17.75% cellulose and 16.28% hemicellulose, while CC consisted of 22.51% lignin, 23.58% cellulose and 33.34% hemicellulose. Alkaline pretreatment was more effective in pretreating CC biomass compared to the SSB biomass.
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- Date Issued: 2020