This week, we characterized lycopene production in E. coli with the pAC-LYC and pAC-LYCipi plasmids, as well as beta-carotene production with the pAC-BETAipi plasmid.
Centrifuging and pelleting overnight cultures of cells containing the pAC-LYC, pAC-LYCipi, and pAC-BETAipi plasmids, in order to isolate the pigment produced.
Measuring the absorption spectra of the pigment.
Resulting spectra.
discussion questions
Using tools such as Biocyc (https://www.biocyc.org) and KEGG (http://www.kegg.jp), review enzymes that can be added to (or knocked out of) your lycopene-producing E. coli to increase the amount of lycopene they produce. Compare the expected and experimental differences in production efficiency between bacteria that carry the pAC-LYC and pAC-LYCipi plasmids.
- Some enzymes upstream of lycopene production in the carotenoid biosythesis pathway map, which can be added to increase production, include:
- crtI - phytoene desaturase
- crtQ - zeta-carotene desaturase
- crtH - prolycopene isomerase
Using Biocyc and/or KEGG, can you identify the enzyme to convert the carrot-orange pigment beta-carotene to the corn-yellow pigment zeaxanthin?
Measuring the absorption spectra of the pigment.
Resulting spectra.