Efficient Expression of Membrane-Bound Water Channel Protein (Aquaporin Z) in Escherichia coli

Jiazhang      Lian; Xiangming      Fang; Jin      Cai; Qixing      Chen; Qiang      Zheng; Lei      Kai; Zhinan      Xu

doi:10.2174/092986608785133717

Abstract

In order to explore the possibility of preparing a high-efficiency aquaporin-based biofilter, an efficient approach for expression of membrane-bound Aquaporin Z (AqpZ) in E. coli was proposed. The AqpZ gene was amplified by means of PCR, and two expression vectors (pET28-AqpZ and pET32-AqpZ) were constructed. The channel protein of interest was synthesized in E. coli BL21(DE3)/pET32-AqpZ as an insoluble fusion protein linked with trxA. However, with BL21(DE3)/pET28-AqpZ, significant amount of AqpZ fused only with 6-His (6-His-AqpZ) could be expressed, correctly folded and targeted into the membrane. Under the optimized culture conditions, the highest expression level (9.05 mg/l) of membrane-bound 6-His-AqpZ was achieved with BL21(DE3)/pET28-AqpZ, and an additional amount (2.35 mg/l) was expressed concomitantly as the inclusion body form. This expression result was 3.5 times higher than that in the previous studies.

Keywords: Water channel protein, Aquaporin Z, Escherichia coli, Membrane-bound expression

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