Journal
JOURNAL OF BIOTECHNOLOGY
Volume 302, Issue -, Pages 85-91Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.jbiotec.2019.06.302
Keywords
L-theanine; Gamma glutamyl transpeptidase; PrsA lipoproteins; Poly(A/T) tails
Categories
Funding
- National Natural Science Foundation of China [21778024]
- National Key Research and Development Program of China [2018YFA0900304]
- China Postdoctoral Science Foundation [2017M620189]
- National First-Class Discipline Program of Light Industry Technology and Engineering [LITE 2018-06]
- Program of Introducing Talents of Discipline to Universities [111-2-06]
- Priority Academic Program Development of Jiangsu Higher Education Institution
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L-theanine, an amino acid known for its favourable taste and linked with health benefits, can be prepared by enzymatic synthesis using gamma-glutamyltranspeptidase (GGT; E.C 2.3.2.2). In the present study, a novel GGT from Bacillus pumilus ML413 was expressed in Bacillus subtlis 168 and exhibited high stability at low temperature (40 degrees C) and alkaline pH 10, compared to the other GGTs. To enhance GGT production, firstly, PrsA lipoproteins was overexpressed in the host which resulted in the extracellular GGT activity doubled. Subsequently, a suitable poly (A/T) tail (TTTAAA) was selected and added to the 3'-terminal of ggt gene, which increased the mRNA stability of ggt gene by 58% and the activity of GGT by 60%. Finally, under optimized fed batch system, L-theanine yield was 53 g l(-1) within 16 h. In this study, we demonstrate a convenient strategy of increasing theanine yield using GGT overexpressing in a safe host B. subtilis 168.
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