重组大肠杆菌全细胞催化合成4-羟基异亮氨酸

4-羟基异亮氨酸(4-HIL)是一种很有前景的药物,它具有促进胰岛素分泌、改善外周组织对胰岛素的抵抗性和调节血脂异常等作用,而L-异亮氨酸羟化酶(IDO)常用于4-HIL的生产。首先,克隆了苏云金芽孢杆菌来源的L-异亮氨酸羟化酶,实现了其在E.coli BL21(DE3)中的异源表达。其次,通过同源建模和蛋白质结构分析,本着将与底物氨基酸侧链结合的氨基酸残基从亲水性或长链疏水性结构突变成丙氨酸Ala的原则,对I156位点进行了定点突变,以增大底物结合口袋,扩宽底物通遒进而提高4-HIL的产量。最后,对野生酶及突变酶的酶学性质、突变酶的羟基化反应体系进行研究,在最优催化条件下,分批补料转化底物进行4-HIL的生产。酶学性质结果显示,野生酶及突变酶I156A的最适温度均为25℃,最适pH均为7.0;突变酶I156A比酶活比野生酶提高了1.9倍,L-ILe转化率提高了28%。羟基化反应体系的最优转化条件为:20 mmol/L L-ILe,20 mmol/Lα-酮戊二酸,8 mmol/L Fe^(2+),30 mmol/L抗坏血酸和HEPES(50 mmol/L,pH 7.0)缓冲液。在最优转化条件下,重组菌E.coli BL21/pET28a-ido^(I156A)进行分批补料转化底物,时间间隔为4 h,32 h后得到77.3 mmol/L 4-HIL,底物最高转化率98.35%e。

Molecular mechanism of different viruses associated with autoimmunity

Different kinds of human chronic diseases may develop the mechanism of autoimmune diseases.As a group of disorders,in the Western world autoimmunity possess as the third most prevalent morbidity and mortality.However,the mechanism of most autoimmune diseases is still under investigation.Viral infection is the principal factor involved in the induction of autoimmune diseases other than genetic factors and cytokine activity.Different mechanisms have been proposed by which viral infection might interrupt tolerance to self and induce autoimmune cascade which eventually leads to the destruction of a specific type of cell or a whole-body organ.The autoimmune attack can be understood through the different immune systems and other possible mechanisms such as molecular mimicry,bystander activation and epitope spreading.In addition to genetic and viral factors,other environmental factors are also involved including bacterial,parasitic and fungal infections.However,different animal models have been studied which provide strong evidence that viruses induced AIDs as well as accelerated and increased lesions in conditions where self-tolerance is interrupted.In the current review,we discussed the virus-induced autoimmunity and the molecular mechanism which is associated with this phenomenon.Here we also discussed the different viruses such as rubella virus,enteroviruses,measles virus,human T-lymphotropic virus type1,human cytomegalovirus,human herpes virus-6,Epstein-Barr virus,rotavirus and some other viruses which modulate the development of AIDs.