Part:BBa_K2092002

PalcA, improved alcR inducible promoter from A. nidulans

The alcA promoter, PalcA, is originally found in Aspergillus nidulans as a part of the ethanol regulon. PalcA requires its positive transcriptional regulator AlcR to regulate the expression of gene alcA. Gene alcA encodes for alcohol dehydrogenase I (ADHI) which facilitates the interconversion between alcohols and carbonyls.

Usage and Biology

PalcA is one of the strongest inducible promoters in Aspergillus nidulans commonly used to overexpress proteins [1]. It has been shown that alcA promoter is also functional in monocotyledonous plant sugar cane [2] and Escherichia coli [3]. Its transciptional activation is dependent on the binding of its positive transcriptional regulator AlcR with various substrates that employ a hydroxyl group, for example ethanol and threonine. The native PalcA consists of 3 AlcR binding sites. The number and position of the AlcR binding sites on the PalcA are crucial in determining its transcriptional activation strength. It has also been shown that each AlcR target in the PalcA contributes differently to the activation of the downstream protein expression.

This part is an improved BioBrick previously characterised by 2011 iGEM DTU-Denmark. Sequencing results showed that two Prefix/Suffix restriction sites XbaI and SpeI were absent in the original part. Hence the two missing restriction sites were added using PCR.

References:

[1] Panozzo, C., Capuano, V., Fillinger, S. and Felenbok, B. (1997). The zinc binuclear cluster Activator AlcR is able to bind to single sites but requires multiple repeated sites for synergistic activation of the alcA gene in Aspergillus nidulans. Journal of Biological Chemistry, 272(36), 22859–22865.

[2] Kinkema, M., Geijskes, R.J., Shand, K., Coleman, H.D., De Lucca, P.C., Palupe, A., Harrison, M.D., Jepson, I., Dale, J.L. and Sainz, M.B. (2013). An improved chemically inducible gene switch that functions in the monocotyledonous plant sugar cane. Plant Molecular Biology, 84(4-5), 443–454.

[3] Hemmati, H. and Basu, C. (2015). Transcriptional analyses of an ethanol inducible promoter in Escherichia coli and tobacco for production of cellulase and green fluorescent protein. Biotechnology & Biotechnological Equipment, 29(6), 1043–1052.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 274
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]