期刊
JOURNAL OF INDUSTRIAL MICROBIOLOGY & BIOTECHNOLOGY
卷 46, 期 8, 页码 1081-1090出版社
SPRINGER HEIDELBERG
DOI: 10.1007/s10295-019-02200-7
关键词
(R)-3; 4-Dihydroxyphenyllactic acid; (S)-3; 4-Dihydroxyphenyllactic acid; 3; 4-Dihydroxyphenyl-l-alanine; Self-sufficient; Whole-cell biotransformation
资金
- National Key Scientific Instrument and Equipment Development Project of China [2013YQ17052504]
- national first-class discipline program of Light Industry Technology and Engineering [LITE2108-04]
- Postgraduate Research & Practice Innovation Program of Jiangsu Province [KYCX19_1841]
- Program for Changjiang Scholars and Innovative Research Team in the University of Ministry of Education of China [IRT_15R55]
- seventh group of Hundred-Talent Program of Shanxi Province (2015)
- Key Project of Research and Development Plan of Shaanxi [2017ZDCXLSF-01-02-01]
We developed an efficient multi-enzyme cascade reaction to produce (R)- or (S)-3,4-Dihydroxyphenyllactic acid [(R)- or (S)-Danshensu, (R)- or (S)-DSS] from 3,4-Dihydroxyphenyl-l-alanine (l-DOPA) in Escherichia coli by introducing tyrosine aminotransferase (tyrB), glutamate dehydrogenase (cdgdh) and d-aromatic lactate dehydrogenase (csldhD) or l-aromatic lactate dehydrogenase (tcldhL). First, the genes in the pathway were overexpressed and fine-tuned for (R)- or (S)-DSS production. The resulting strain, E. coli TGL 2.1 and E. coli TGL 2.2, which overexpressed tyrB with the stronger T7 promoter and cdgdh, csldhD or tcldhL with the weaker Trc promoter, E. coli TGL 2.1 yielded 57% increase in (R)-DSS production: 59.8 +/- 2.9 mM. Meanwhile, E. coli TGL 2.2 yielded 54% increase in (S)-DSS production: 52.2 +/- 2.4 mM. The optimal concentration of L-glutamate was found to be 20 mM for production of (R)- or (S)-DSS. Finally, l-DOPA were transformed into (R)- or (S)-DSS with an excellent enantiopure form (enantiomeric excess > 99.99%) and productivity of 6.61 mM/h and 4.48 mM/h, respectively.
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