Types | DnaRegion
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Roles | Generator
engineered_region
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Sequences | BBa_K814012_sequence (Version 1)
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Description
The induction of this novel pathway in S. cerevisiae requires two additional enzymes from Coffea canephora: XMT1 and DXMT1. S. cerevisiae will produce the required metabolic precursors to Xanthosine, where after the XMT1 enzyme from C. canephora will use the substrate to synthesize 7-methylxanthosine. DXMT1 will continue the synthesis by converting to 7-Methylxanthine, where in XMT1 will again convert it to Theobromine. Finally DXMT1 will convert the metabolite to caffeine. Both the XMT1 and DXMT1 genes produce bifunctional enzymes and this particular pathway was chosen so that one less enzyme would be required in the synthesis (compared to three in C. arabica).
This part includes the yeast pCyc promoter, a yeast Kozak sequence, the Coffea canephora XMT1 open reading frame which has been codon-optimized for yeast, and the tCycE1 terminator. It can be used to generate XMTI protein in yeast.
Notes
The sequences generated by the "de-optimization" software were ordered from IDT as G-Blocks and assembled using overlap-extension PCR.
Source
The sequences were generated using a program developed by our team which takes multiple codon-optimized sequences, looks for regions of homology between them, and disrupts these regions by ???de-optimizing??? the region in such a way as to eliminate homology. This is important for developing and assembling yeast BioBrickTM parts in such a way that designed sequences will not undergo homologous recombination.