Journal
APPLIED MICROBIOLOGY AND BIOTECHNOLOGY
Volume 89, Issue 5, Pages 1387-1394Publisher
SPRINGER
DOI: 10.1007/s00253-010-2994-4
Keywords
Tryptophan decarboxylase; Tryptamine 5-hydroxylase; Dual expression; Serotonin; Escherichia coli
Categories
Funding
- Ministry of Agriculture and Forestry [108078-3]
- Rural Development Administration [20100301-061-023-001-03-00]
- Ministry of Education, Science, and Technology [2010-0020141]
- Institute of Planning & Evaluation for Technology in Food, Agriculture, Forestry & Fisheries (iPET), Republic of Korea [IPET108078-3] Funding Source: Korea Institute of Science & Technology Information (KISTI), National Science & Technology Information Service (NTIS)
- National Research Foundation of Korea [2010-0020141] Funding Source: Korea Institute of Science & Technology Information (KISTI), National Science & Technology Information Service (NTIS)
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A plant-specific biogenic amine, serotonin, was produced by heterologous expression of two key biosynthetic genes, tryptophan decarboxylase (TDC) and tryptamine 5-hydroxylase (T5H), in Escherichia coli. The native T5H, a cytochrome P450 enzyme, was unable to be functionally expressed in E. coli. Through a series of N-terminal deletions or additions of tagging proteins, we generated a functional T5H enzyme construct (GSTa dagger 37T5H) in which glutathione S transferase (GST) was translationally fused with the N-terminal 37 amino acid deleted T5H. Dual expression of GSTa dagger 37T5H and TDC using a pCOLADuet-1 E. coli vector produced serotonin at concentrations of approximately 24 mg l(-1) in the culture medium and 4 mg l(-1) in the cells. An optimum temperature of approximately 20A degrees C was required to achieve peak serotonin production in E. coli because the low induction temperature gave rise to the highest soluble expression of GSTa dagger 37T5H.
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