Abstract
Two thermophilic 1,4-α-glucan branching enzymes (GBEs), CbGBE from Caldicellulosiruptor bescii and PhGBE from Pyrococcus horikoshii, which belong to the glycoside hydrolase family 13 and 57 respectively, were cloned and expressed in Escherichia coli. Two GBEs were identified to have α-1,6 branching activity against various substrates, but substrate specificity was distinct. Starch was modified by two GBEs and their in vitro digestibility and structural properties were investigated. Short-branched A chains with a degree of polymerization (DP) of 6–12 increased with CbGBE-modified starch, increasing the proportion of slow digestible and resistant starch (RS) fractions. PhGBE-modified starch resulted in an increase in the RS fraction only by a slight increase in part of A chains (DP, 6–9). Compared to the proportion of control not treated with GBE, the proportion of α-1,6 linkages in CbGBE- and PhGBE-modified starch increased by 3.1 and 1.6 times. 13C cross polarization/magic angle sample spinning (CP/MAS) NMR and XRD pattern analysis described that GBE-modified starches reconstructed double helices but not the crystalline structure. Taken together, CbGBE and PhGBE showed distinct branching activities, resulting in different α-1,6 branching ratios and chain length distribution, and double helices amount of starch, ultimately affecting starch digestibility. Therefore, these GBEs can be used to produce customized starches with controlled digestion rates.
Highlights
Starch is the most abundant and important polysaccharide consumed as an energy source by humans
The PhGBE from P. horikoshii OT3 and CbGBE from C. bescii DSM 6725 were expressed in E. coli
Two thermophilic enzymes belonging to two different families, CbGBE (GH13) and PhGBE
Summary
Starch is the most abundant and important polysaccharide consumed as an energy source by humans. Starch is divided into three categories based on its rate of digestion to glucose units in the small intestine: rapidly digestible starch (RDS), slowly digestible starch (SDS), and resistant starch (RS) [1]. SDS is slowly but completely digested in the small intestine, leading to a gradual increase in postprandial blood glucose levels. RS is not digested in the small intestine. It moves to the large intestine, where it is fermented by the intestinal microflora [2]. SDS and RS have low glycemic index (GI) values. The consumption of low-GI foods has health benefits that include a reduced risk of chronic metabolic diseases, such as diabetes, obesity, and cardiovascular disease [3]
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