Effects of barley and oat ß-glucan structures on their rheological and thermal characteristics.

In order to understand the relationship between chemical structure and physical properties of cereal ß-glucans, the ß-glucans with identical Mw (98.4-99.2 kDa) and Rg (21.1-22.0 nm) were isolated from chal and gwangan barley, and ohl oat, and their linkage structure, flow behavior, and thermal properties were investigated. Previously, we established a purification method of 3-O-cellobiosyl-glucose (DP3) and 3-O-cellotriosyl-glucose (DP4) (Yoo, Lee, Chang, Lee, & Yoo, 2007) and applied these authentic standards to quantify the ratio of ß-(1,4)/(1,3) linkages in cereal ß-glucans. ß-Glucans isolated from two barley cultivars had greater proportion of DP3 than did the oat, and within barley cultivars chal barley ß-glucan had significantly larger amount of DP3 over gwangan cultivar. Thus, chal barley ß-glucan had the greatest molar ratio (2.53) of DP3 to DP4, and ohl oat had the lowest (1.51). While all the ß-glucan solutions showed strong shear thinning behavior, ohl oat ß-glucan with higher proportion of DP4 exhibited the highest viscosity among the ß-glucan samples. After 3 freeze-thaw cycles of 3% (w/v) ß-glucan samples, chal barley ß-glucan had lower onset (To) and peak (Tp) temperatures (28.3 and 36.7 °C, respectively) than those of gwangan barley (33.6 and 39.9 °C) and ohl oat (37.9 and 46.9 °C) did, and the heat scan profiles were thermoreversible. The To and Tp of inter-chain associations decreased as the DP3:DP4 ratio of the ß-glucan increased. From this study, it was suggested that cellotetraosyl units and longer ß-(1,4)-linked segments would be a major contributor for improving solution viscosity and gel formation of cereal ß-glucans.

Effect of amylose content in corn starch modification by Thermus aquatiqus 4-alpha-glucanotransferase.

Corn starches with different amylose contents were enzymatically modified using Thermus aquaticus 4-alpha-glucanotransferase (TAalphaGTase). On treating, chain length distribution of the product became broader (degree of polymerization; DP 3-40) after isoamylolysis when compared to the untreated corn starch. In addition, a variety of different size cycloamyloses were formed by glucanotransferring activity of TAalphaGTase. Cycloamyloses (CAs) with DP 5-40 were detectable in all of the TAalphaGTase-treated corn starches. The amount of CAs produced by the enzyme treatment seems to increase as amylose content of starch increased when judged by high-performance anion exchange chromatography (HPAEC) and high-performance size exclusion chromatography (HPSEC) analyses. Thus, it was suggested that the extent of modification on starch molecules was enhanced in proportion to amylose content of starch by transferring activity of TAalphaGTase. In turn, it could be useful for developing an efficient process of CA production using this enzyme.

Comparative structural analyses of purified glycogen particles from rat liver, human skeletal muscle and commercial preparations.

Glycogen is a cellular energy store that is crucial for whole body energy metabolism, metabolic regulation and exercise performance. To understand glycogen structure we have purified glycogen particles from rat liver and human skeletal muscle tissues and compared their biophysical properties with those found in commercial glycogen preparations. Ultrastructural analysis of commercial liver glycogens fails to reveal the classical alpha-rosette structure but small irregularly shaped particles. In contrast, commercial slipper limpet glycogen consists of beta-particles with similar branching and chain lengths to purified rat liver glycogen together with a tendency to form small alpha-particles, and suggest it should be used as a source of glycogen for all future studies requiring a substitute for mammalian liver glycogen.

Heterologous expression and characterization of glycogen branching enzyme from Synechocystis sp. PCC6803.

A gene (sll0158) putatively encoding a glycogen branching enzyme (GBE, E.C. 2.4.1.18) was cloned from Synechocystis sp. PCC6803, and the recombinant protein expressed and characterized. The PCR-amplified putative GBE gene was ligated into a pET-21a plasmid vector harboring a T7 promoter, and the recombinant DNA transformed into a host cell, E. coli BL21(DE3). The IPTG-induced enzymes were then extracted and purified using Ni-NTA affinity chromatography. The putative GBE gene was found to be composed of 2,310 nucleotides and encoded 770 amino acids, corresponding to approx. 90.7 kDa, as confirmed by SDS-PAGE and MALDI-TOF-MS analyses. The optimal conditions for GBE activity were investigated by measuring the absorbance change in iodine affinity, and shown to be pH 8.0 and 30 degrees in a 50 mM glycine-NaOH buffer. The action pattern of the GBE on amylose, an alpha-(1,4)-linked linear glucan, was analyzed using high-performance anion-exchange chromatography (HPAEC) after isoamylolysis. As a result, the GBE displayed alpha-glucosyl transferring activity by cleaving the alpha-(1,4)-linkages and transferring the cleaved maltoglycosyl moiety to form new alpha-(1,6)- branch linkages. A time-course study of the GBE reaction was carried out with biosynthetic amylose (BSAM; Mp approximately = 8,000), and the changes in the branch-chain length distribution were evaluated. When increasing the reaction time up to 48 h, the weight- and number-average DP (DPw and DPn) decreased from 19.6 to 8.7 and from 17.6 to 7.8, respectively. The molecular size (Mp, peak Mw approximately = 2.45-2.75 x 10(5)) of the GBE-reacted product from BSAM reached the size of amylose (AM) in botanical starch, yet the product was highly soluble and stable in water, unlike AM molecules. Thus, GBE-generated products can provide new food and non-food applications, owing to their unique physical properties.