A glycogen branching enzyme from Thermomonospora curvata: Characterization and its action on maize starch

Abstract

A gene (TcGBE) encoding a glycogen branching enzyme from Thermomonospora curvata was expressed in Escherichia coli and purified, and biochemically analyzed. The purified enzyme was approximately 84 kDa, and exhibited a maximum activity at 55°C and pH 8.5. The enzyme showed the highest specificity to amylose with a specific activity of 90.28 U/mg. The recombinant enzyme could act on maize starch, and starch treated with TcGBE was observed to have a lower amylose content and molecular weight. The number of short chains of amylopectin in TcGBEmodified starch with a degree of polymerization (DP) less than 13 increased from 20.19 to 32.59%, and was accompanied by a reduction in longer chains (DP > 25) from 25.58 to 10.03%. In addition, the medium length chains of amylopectin (13 < DP <25) also showed a slight increase from 54.23 to 58.38%. The 1 H NMR spectra revealed an increased ratio of a-1,6-to a-1, 4-glucosidic linkages from 4.1 to 10.3% following treatment with TcGBE for 10 h. Taken together, these results suggested that TcGBE could be used to generate a modified maize starch with a low amylose content and more branched amylopectin of low molecular weight but richer in short and intermediate chains.

Synthesis of highly branched α-glucans with different structures using GH13 and GH57 glycogen branching enzymes

“…However, many more bacterial GH13 GBEs have been characterized, among which the GBEs of Aquifex aeolicus (Hiroki Takata, Ohdan, Takaha, Kuriki, & Okada, 2003;M. Van Der Maarel, Vos, Sanders, & Dijkhuizen, 2003), Butyrivibrio fibrisolvens (Rumbak, Rawlings, Lindsey, & Woods, 1991), Deinococcus geothermalis and Deinococcus radiodurans (Palomo, Kralj, van der Maarel, & Dijkhuizen, 2009), Geobacillus stearothermophilus (Hiroki Takata et al, 1994), Geobacillus thermoglucosidans (Ban et al, 2016;Liu et al, 2017), Mycobacterium tuberculosis (Garg, Alam, Kishan, & Agrawal, 2007), Thermomonospora curvata (Fan, Xie, Zhan, Chen, & Tian, 2016), and Vibrio vulnificus (Jo, Park, Jeong, Kim, & Park, 2015). Today only three GH57 GBEs have been characterized; Pyrococcus horikoshii (Na et al, 2017), Thermococcus kodakarensis (Murakami, Kanai, Takata, Kuriki, & Imanaka, 2006), and Thermus thermophilus (Palomo et al, 2011).…”

Glycogen Branching Enzyme with a Novel Chain Transfer Mode Derived from Corallococcus sp. Strain EGB and Its Potential Applications

“…The λ max and A λ max values of iodine–starch complexes are correlated with the degree of polymerization (DP) of amylose and the CL of amylopectin . The formation of iodine complexes demands a helical conformation of chains with at least six d -glucose residues in one helix turn. , This indicates that both the longer internal or external chains of amylopectin and amylose can combine with iodine to form a complex.…”

“…One unit of enzyme activity (U) was defined as the amount of enzyme required to decrease the absorbance value by 1% per min. 21 The absorbance spectra of the reaction mixtures treated for different times were analyzed by the scanning method over the wavelength range of 450−800 nm.…”

“…Two hundred microliters of reaction mixture was added to standard 96-well plates, and the absorbance was measured at 660 nm ( A 660 ) using a SpectraMax i3x (Molecular Devices, USA). One unit of enzyme activity (U) was defined as the amount of enzyme required to decrease the absorbance value by 1% per min . The absorbance spectra of the reaction mixtures treated for different times were analyzed by the scanning method over the wavelength range of 450–800 nm.…”