Shibasaki T, Hashimoto S, Mori H, Ozaki A
Development Department, Bio-chemicals Company, Kyowa Hakko Kogyo Co. Ltd., 6-1 Ohtemachi, 1-chome, Chiyoda-ku, Tokyo 100-8185, Japan.
J Biosci Bioeng. 2000;90(5):522-5. doi: 10.1016/s1389-1723(01)80033-5.
An Escherichia coli recombinant strain producing trans-4-hydroxy-L-proline (Hyp) was constructed by introducing a proline 4-hydroxylase gene into an L-proline-producing E. coli. Plasmid pPF1, which contains a gene encoding feedback resistant gamma-glutamyl kinase (proB74), was constructed and introduced into E. coli W1485 putA. The recombinant E. coli W1485 putA/pPF1 strain produced L-proline (1.2 g/l). The proline production by W1485 putA/pPF1 was converted to Hyp production by introducing pWFH1 which contains a proline 4-hydroxylase gene. E. coli W1485 putA which harbors pWFP1 carrying the proline 4-hydroxylase gene, proB74, and proA produced 25 g/l of Hyp in 96 h. A novel biosynthetic pathway of Hyp, which has not previously been produced in E. coli, was constructed in E. coli.
通过将脯氨酸4-羟化酶基因导入产L-脯氨酸的大肠杆菌中，构建了一株产反式-4-羟基-L-脯氨酸（Hyp）的大肠杆菌重组菌株。构建了含有编码反馈抗性γ-谷氨酰激酶（proB74）基因的质粒pPF1，并将其导入大肠杆菌W1485 putA中。重组大肠杆菌W1485 putA/pPF1菌株产L-脯氨酸（1.2 g/l）。通过导入含有脯氨酸4-羟化酶基因的pWFH1，将W1485 putA/pPF1的脯氨酸产量转化为Hyp产量。携带脯氨酸4-羟化酶基因、proB74和proA的pWFP1的大肠杆菌W1485 putA在96小时内产25 g/l的Hyp。在大肠杆菌中构建了一条此前未在大肠杆菌中产生的Hyp新生物合成途径。
