Xylanase gene mutation by error-prone pcr and expression in Pichia pastoris / Mutação do gene da xilanase por pcr propensa a erros e expressão em Pichia pastoris

Xylanase can hydrolyze xylan for reducing its anti-nutritional impact and improving nutrient availability, so obtaining suitable xylanase to degrade xylan is essential. Error-prone PCR and gene transformation were used in this study to obtain the ideal xylanase for degrading xylan effectively. The result showed that one mutant xylanase gene with high xylanase expression was obtained. After the mutant xylanase gene was connected with pGAPZA and transformed into Pichia pastoris (P. pastoris), the recombinant P. pastoris with mutant gene was found to produce higher xylanase activity (0.1480 U/mL) than that with the native xylanase gene (0.1360 U/mL) after 12 h incubation (p<0.05). The optimal temperature and pH of xylanase expressed by native and mutant genes were the same, i.e. 40°C and 5.50 (p<0.05). In addition, adding 0.2% Tween 80 during recombinant P. pastoris incubation could significantly increase xylanase yield by about 30-35% (p<0.05). The mutant xylanase could significantly increase xylose yield from wheat meal more than the native xylanase (p<0.05).

A xilanase pode hidrolisar o xilano para reduzir seu impacto antinutricional e melhorar a disponibilidade de nutrientes, portanto, obter xilanase adequada para degradar o xilano é essencial. A PCR propensa a erros e a transformação genética foram utilizadas neste estudo para obter a xilanase ideal para degradar eficazmente a xilana. O resultado mostrou que um gene mutante de xilanase com alta expressão de xilanase foi obtido. Depois que o gene mutante da xilanase foi conectado ao pGAPZA e transformado em Pichia pastoris (P. pastoris), o recombinante P. pastoris com o gene mutante produziu maior atividade de xilanase (0,1480 U / mL) do que com o gene nativo da xilanase (0,1360 U / mL) após 12 h de incubação (p <0,05). A temperatura e o pH ótimos da xilanase expressa pelos genes nativos e mutantes foram os mesmos, ou seja, 40 ºC e 5,50 (p <0,05). Além disso, a adição de Tween 80 a 0,2% durante a incubação de P. pastoris recombinante poderia aumentar significativamente o rendimento de xilanase em cerca de 30-35% (p <0,05). A xilanase mutante poderia aumentar significativamente o rendimento de xilose da farinha de trigo mais do que a xilanase nativa (p <0,05).

Tea Polyphenols Exerts Anti-hepatitis B Virus Effects in a Stably HBV-transfected Cell Line / 华中科技大学学报（医学）（英德文版）

In this study,the anti-HBV effects of tea polyphenols (TP) were examined.After cells were exposed to TP for 3,6,9 days,amounts of HBsAg,HBeAg and HBV-DNA released into the supernatant of the cultured HepG2 2.2.15 cells were detected.TP,to some extent,inhibited the secre-tion of HBsAg and strongly suppressed the secretion of HBeAg in a dose-dependent (P<0.01) and time-dependent manner,with 50% maximal inhibitory concentration (IC50) value being 7.34 μg/mL on the 9th day,but the time-dependence was not significant (P=0.051).Expression of HBV-DNA in the supernatant of the cell culture also was significantly decreased in a dose-dependent fashion (P<0.01).The IC50 of TP in inhibiting HBV DNA was 2.54 μg/mL.It concluded that TP possessed potential anti-HBV effects and may be used as a treatment alternative for HBV infection.