Modulating Cell Permeability for Whole-Cell Biocatalysis in Chemical Synthesis (TSE03-D)

全细胞生物催化的一个基本问题是细胞膜对各种工业底物的不渗透性，这极大地限制了反应速度，在最糟糕的情况下，使细胞内的酶网络完全无法进入。这项研究针对渗透性问题提出了两种新的策略。在第一种策略中，一种具有脂多糖生物合成修饰的大肠杆菌Lipid A突变体将被用来改变外膜的通透性。在第二种策略中，将使用可以改变膜通透性的膜活性多肽。这些多肽将通过精心设计的选择和筛选程序通过定向进化方法产生。细胞通透性的调节可以通过改变阳离子浓度、通透性多肽的表达水平或使用具有不同膜通透性活性的多肽序列来实现。通过去除渗透性障碍，全细胞生物催化剂可以接触到各种类型的合成底物、疏水和大型亲水底物，这些底物通常没有细胞吸收的机制。

One fundamental problem in whole-cell biocatalysis is the impermeability of cell membranes to diverse substrates of industrial interests, which greatly limits the reaction rate, and in worst scenarios makes the intracellular enzyme network completely inaccessible. This research proposes two novel strategies to the permeability issue. In the first strategy, an E.coli Lipid A mutant with modified biosynthesis of lipopolysaccharides will be used to alter the outer membrane permeability. In the second strategy, membrane-active peptides that modify membrane permeability will be used. These peptides will be generated by directed evolution methods with carefully designed selection and screening procedures. The modulation of cell permeability could be effected by varying cation concentrations, permeabilizing peptide expression level or by using a peptide sequence with a different membrane permeabilizing activity. By removing permeability barriers, the whole-cell biocatalysts can be made accessible to synthetic substrate of various types, hydrophobic and large hydrophilic substrates that cells do not normally have mechanisms for uptake.