Effect of Blueberry Anthocyanins Malvidin and Glycosides on the Antioxidant Properties in Endothelial Cells.

The objective of this research was to survey the antioxidant functional role of the main anthocyanins of blueberries in endothelial cells. Changes on the reactive oxygen species (ROS), xanthine oxidase-1 (XO-1), superoxide dismutase (SOD), and heme oxygenase-1 (HO-1) in cells of malvidin and the two glycosides were investigated. The results showed that these anthocyanins decreased the levels of ROS and XO-1 but increased the levels of SOD and HO-1. Glycosides improved the antioxidant capacity of malvidin to a great extent. The changes in the antioxidant properties of malvidin-3-glucoside were more pronounced than malvidin-3-galactoside. Variation in levels of malvidin-3-glucoside and malvidin-3-galactoside had a significant impact on antioxidant properties to different extents. It indicates that blueberries are a good resource of anthocyanins, which can protect cells from oxidative deterioration and use blueberry as a potential functional food to prevent diseases related to oxidative stress.

[Construction and characterization of R169H mutant of aspartokinase from Corynebacterium pekinense].

OBJECTIVE: Increasing the activity of aspartokinase (AK) from Corynebacterium pekinense. METHODS: The gene of AK was constructed and mutated by site-specific mutagenesis. The mutational recombinant plasmid was heterologously expressed in Escherichia coli BL21. The mutational AK was purified by Ni(2+)-NTA column after ultrasonicating of the recombinant bacteria, and then identified by SDS-PAGE and Western blot. We compared the kinetic difference between R169H AK and WT AK by determining the enzymatic activities. Some other characteristics of R169H AK and WTAK were also studied. RESULTS: The mutant R169H was successfully constructed. The molecular weight of AK was 48kDa. V(max) of R169H AK was 226.3 U/mg x s(-1), which was 2.3 times higher than that of WT AK. The optimum reaction temperature of R169H AK was 26 degrees C, the same as that of WT AK. The optimum reaction pH of R169H AK was 9.0, slightly higher than that of WT AK. The half-life period of R169H AK under optimum temperature and pH were 5.5h, much higher than that of WT AK. Lysine, threonine and methionine had an active effect on the activity of R169H AK when they were in low concentration. CONCLUSION: The hydrogen bond between R169 and E92 was broken down in R169H AK, which could affect the degree of polymerization and further lowered the affinity of mutant AK with substrates and then decreased the inhibition inducing by the metabolites. Thus, the V(max) of mutant AK from R169H had increased by 2.3 times compared with that of WT AK.

[Heterologous expression and characterization of L200F/D215K mutant of homoserine dehydrogenase from Corynebacterium pekinense AS1.299].

OBJECTIVE: To obtain a new homoserine dehydrogenase with better properties from Corynebacterium pekinense by the spatial structure transfromation. METHODS: Double mutants L200F/D215A, L200F/D215E, L200F/D215G and L200F/D215K were constructed by site-directed mutagenesis and expressed in E. coli BL21. L200F/D215K was characterized for its highest catalytic efficiency and compared with that of L200F. RESULTS: The Vmax of L200F/D215K was 36.92 U/mg, 1.24 times as that of L200F. The optimum reaction temperature of L200F/D215K was 37 degrees C, 2 degrees C higher than that of L200F. The optimum pH of L200F/D215K was 7.5, the same as that of L200F. The half-life time of L200F/D215K under optimum temperature was 4.16 h and was 1.12 times as that of L200F. Both L200F/D215K and L200F had good resistance to organic solvents and metal ions. CONCLUSION: Through the spatial structure transformation, the enzymatic activity was increased, and the enzymology properties was optimized.