Production of thermostable endo-1,5-α-l-arabinanase in Pichia pastoris for enzymatically releasing functional oligosaccharides from sugar beet pulp

• Published in: Applied microbiology and biotechnology Vol. 104; no. 4; pp. 1595 - 1607
• Main Authors: Zhang, Ningning, Wright, Tristen, Wang, Xiaoting, Savary, Brett J., Xu, Jianfeng
• Format: Journal Article
• Language: English
• Published: Berlin/Heidelberg Springer Berlin Heidelberg 01.02.2020; Springer Nature B.V
• Subjects: Arabinanase; Arabinose; Biomedical and Life Sciences; Biotechnologically Relevant Enzymes and Proteins; Biotechnology; Cell culture; Cell walls; Colon; Crop residues; Enzymatic activity; Enzyme activity; Enzymes; Epithelial cells; Ferulic acid; Functional foods & nutraceuticals; Glycosidases; Glycoside hydrolase; Hydrolase; Hydrophobic chromatography; Hydrophobicity; Life Sciences; Microbial Genetics and Genomics; Microbiology; Oligosaccharides; Oxidation; Pichia pastoris; Polysaccharides; Proteins; Pulp; Sugar; Thermal stability; Yeast; Yeasts; Zonula occludens-1 protein; Sugar beet pulp; Functional oligosaccharides; Arabinanase; Intestinal epithelium; Tight junction protein; Feruloylated arabino-oligosaccharides; Pichia pastoris
• ISSN: 0175-7598; 1432-0614
• DOI: 10.1007/s00253-019-10238-x

Sugar beet pulp is an agricultural processing residue that is a rich source of the cell wall polysaccharide arabinan. Functional oligosaccharides, specifically feruloylated arabino-oligosaccharides (FAOs), can be isolated from sugar beet pulp through selective action by endo-arabinanase (glycoside hydrolase family 43). This study aimed to develop yeast ( Pichia pastoris ) as an efficient, eukaryotic platform to produce a thermophilic endo-1,5-α- l -arabinanase (TS-ABN) for extracting FAOs from sugar beet pulp. Recombinant TS-ABN was secreted into yeast culture medium at a yield of ~ 80 mg/L, and the protein exhibited specific enzyme activity, pH and temperature optimum, and thermostability comparable to those of the native enzyme. Treatment of sugar beet pulp with Pichia -secreted TS-ABN released FAOs recovered by hydrophobic chromatography at 1.52% (w/w). The isolated FAOs averaged seven arabinose residues per ferulic acid, and treatment of T84 human colon epithelial cells significantly increased expression of two key tight junction-related proteins—zonula occludens-1 and occludin—in a dose-dependent manner. This research establishes a biochemical platform for utilizing sugar beet pulp to produce value-added bioproducts with potential nutraceutical applications.