School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, PR China; National Engineering Laboratory for Cereal Fermentation Technology, School of Biotechnology, Jiangnan University, Wuxi 214122, PR China.
School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, PR China.
Bioresour Technol. 2017 Nov;244(Pt 1):1104-1110. doi: 10.1016/j.biortech.2017.08.039. Epub 2017 Aug 10.
The objective of the study was to enhance the substrate tolerance of Pseudomonas putida nitrilase via atmospheric and room temperature plasma (ARTP) and cell immobilization. The mutant library was constructed by ARTP and rapidly screened by an OPA-TCA microscale reaction. A mutant strain of mut-D3 was obtained and its optimum substrate concentration was improved to 150mM from 100mM. It could accumulate 189g/L nicotinic acid (NA) from 3-cyanopyridine (3-CP), which was increased by 42% compared with that of wild type (WT). Additionally, composite immobilization of mut-D3 was performed and SA-PVA immobilized cells could catalyze 250mM 3-CP each batch with finally accumulating 346g/L NA, while free cells accumulated 175g/L NA. These results indicated that the free or immobilized catalysts of mut-D3 could serve as a good choice for NA production. This is the first report on mutation breeding of nitrilase-producing microorganisms by ARTP.
本研究旨在通过大气压室温等离子体（ARTP）和细胞固定化提高假单胞菌腈水解酶的底物耐受性。通过 ARTP 构建突变文库，并通过OPA-TCA 微量反应快速筛选。获得了突变株 mut-D3，其最佳底物浓度从 100mM 提高到 150mM。与野生型（WT）相比，它可以从 3-氰基吡啶（3-CP）中积累 189g/L 烟酸（NA），增加了 42%。此外，还进行了 mut-D3 的复合固定化，SA-PVA 固定化细胞每批可催化 250mM 3-CP，最终积累 346g/L NA，而游离细胞积累 175g/L NA。这些结果表明，mut-D3 的游离或固定化催化剂可以作为生产 NA 的良好选择。这是首次报道通过 ARTP 对产腈酶微生物进行诱变育种。
