Partitioning and purification of extracellular β-1,3-1,4-glucanase in aqueous two-phase systems

INTRODUCTION β-1,3-1,4-glucan is the major cell wall compo-nent of barley and other cereal endosperms. It consistsof a polymer of glucose monomers joined by β-1,3-and β-1,4-glycosidic bonds in an irregular fashion(Woodward et al., 1983; Edney et al., 1991). Due toits unique structure, barley β-glucan can be dissolvedin water in high molecular mass and resulting in ahigh viscosity solution. The high-molecular-massβ-glucan released during mashing causes severe fil-tration problems and…     

A food-grade industrial arming yeast expressing β-1,3-1,4-glucanase with enhanced thermal stability

The aim of this work was to construct a novel food-grade industrial arming yeast displaying β-1,3-1,4-glucanase and to evaluate the thermal stability of the glucanase for practical application. For this purpose, a bi-directional vector containing galactokinase (GAL1) and phosphoglycerate kinase 1 (PGK1) promoters in different orientations was constructed. The β-1,3-1,4-glucanase gene from Bacillus subtilis was fused to α-agglutinin and expressed under the control of the GAL1 promoter. α-galactosidase induced by the constitutive PGK1 promoter was used as a food-grade selection marker. The feasibility of the α-galactosidase marker was confirmed by the growth of transformants harboring the constructed vector on a medium containing melibiose as a sole carbon source, and by the clear halo around the transformants in Congo-red plates owing to the expression of β-1,3-1,4-glucanase. The analysis of β-1,3-1,4-glucanase activity in cell pellets and in the supernatant of the recombinant yeast strain revealed that β-1,3-1,4-glucanase was successfully displayed on the cell surface of the yeast. The displayed β-1,3-1,4-glucanase activity in the recombinant yeast cells increased immediately after the addition of galactose and reached 45.1 U/ml after 32-h induction. The thermal stability of β-1,3-1,4-glucanase displayed in the recombinant yeast cells was enhanced compared with the free enzyme. These results suggest that the constructed food-grade yeast has the potential to improve the brewing properties of beer.     

Partitioning and purification of extracellular β-1,3-1,4-glucanase in aqueous two-phase systems

The partition behaviors of β-1,3-1,4-glucanase, α-amylase and neutral proteases from clarified and whole fermentation broths of Bacillus subtilis ZJF-1A5 were investigated. An aqueous two-phase system (polyethylene glycol (PEG)/MgSO4)was examined with regard to the effects of PEG molecular weight (MW) and concentration, MgSO4 concentration, pH and NaCl concentration on enzyme partition and extraction. The MW and concentration of PEG were found to have significant effects on enzyme partition and extraction with low MW PEG showing the greatest benefit in the partition and extraction of β-glucanase with the PEG/MgSO4 system. MgSO4 concentration influenced the partition and extraction of β-glucanase significantly. pH had little effect on β-glucanase or proteases partition but affected α-amylase partition when pH was over 7.0. The addition of NaCl had little effect on the partition behavior of β-glucanase but had very significant effects on the partitioning of α-amylase and on the neutral proteases. The partition behaviors of β-glucanase, α-amylase and proteases in whole broth were also investigated and results were similar to those obtained with clarified fermentation broth. A two-step process for purifying β-glucanase was developed, which achieved β-glucanase recovery of 65.3% and specific activity of 14027 U/mg, 6.6 times improvement over the whole broth.     

Optimization of cultural conditions for thermostable β-1,3-1,4-glucanase production by Bacillus subtilis ZJF-1A5

INTRODUCTIONβ 1 ,3 1 ,4 glucanisthemajorcellwallcom ponentofbarleyandothercerealendosperm ,andconsistsofpolymersofglucosemonomersjoinedbyβ 1 ,3 andβ 1 ,4glycosidicbondsinanon regularfashion (Woodwardetal.,1 983 ;Edneyetal.,1 991 ) .High molecular mas…     

1,4-二氢吡啶衍生物与DNA相互作用的荧光光谱研究

用荧光光谱研究了1,4-二氢吡啶衍生物(1,4-DHP)与小牛胸腺DNA(CT-DNA)的相互作用。结果表明,1,4-DHP衍生物分子与CT-DNA相互作用后,如其荧光显著增强,表明1,4-DHP衍生物分子嵌入到CT-DNA中,如荧光明显减弱,表明1,4-DHP衍生物分子不是以嵌入模式与CT-DNA相互作用。     

(E)-2-(1,3-二硫戊环-2-亚基)-3-羰基-5-芳基-4-戊烯酸酯的合成

成功地实现了对酸敏感的（E）-2-（1,3-二硫戊环-2-亚基）-3-羰基-5-芳基4-戊烯酸1与脂肪醇的酯化反应,以较好的产率生成相应的酯3．