Part:BBa_K4588038
Contents
4-hydroxy phenylacetate 3-monooxygenase reductase component B (S. cerevisiae), HpaB
This part encodes 4-hydroxy phenylacetate 3-monooxygenase reductase component B from Lysinibacillus capsici
Biology
The organism this gene is initially expressed in is L. capsici.
HpaB is a reduced FAD (FADH2)-utilizing monooxygenase. This enzyme component uses FADH2 and O2 to oxidize 4-hydroxyphenylacetate [1].
Design
This BioBrick contains a GAL1 promoter (BBa_K2637059) that supports strong inducible expression in the presence of galactose. Following this promoter, the sequence contains a Kozak sequence (BBa_J63003) for optimal ribosome binding. The BioBrick for HpaB consists of the coding region HpaB (BBa_K4588027) and is codon optimized for S. cerevisiae expression. Following the coding regions, a 6x His Tag (BBa_K1223006) was inserted for protein expression detection. The strong ADH1 terminator (BBa_K1486025) was chosen to terminate gene transcription completely.
Usage
This enzyme is implemented in the synthesis pathway to produce rosmarinic acid in the Saccharomyces cerevisiae culture. HpaB is part of the two-step process of converting p-coumaric acid into caffeic acid, which adds a hydroxyl group onto the phenyl ring.
References
1. Xun, L., & Sandvik, E. R. (2000). Characterization of 4-hydroxyphenylacetate 3-hydroxylase (HpaB) of Escherichia coli as a reduced flavin adenine dinucleotide-utilizing monooxygenase. Applied and environmental microbiology, 66(2), 481–486. https://doi.org/10.1128/AEM.66.2.481-486.2000
2. Babaei, M., Borja Zamfir, G. M., Chen, X., Christensen, H. B., Kristensen, M., Nielsen, J., & Borodina, I. (2020). Metabolic engineering of saccharomyces cerevisiae for rosmarinic acid production. ACS Synthetic Biology, 9(8), 1978–1988. https://doi.org/10.1021/acssynbio.0c0004
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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