Part:BBa_K2310100:Design
LuxAB, emitting luciferase (from X. luminescens)
Luminous bacteria are the most abundant and widely distributed of the light-emitting organisms and are found in marine, freshwater, and terrestrial environments. What their most important feature is they can produce the luciferase called LUXAB, which can catalyzes the bioluminescence reactions. Almost all luminous bacteria have been classified into the three genera Vibrio, Photobacterium, and Xenorhabdus.In our progress, the luciferase what we use is from the Xenorhabdus luminescens.
LuxAB is a part of luxCDABEG which is the normal structure of the operon in most bioluminescent bacteria. The LuxCDE gene controls the synthesis/regenerate aldehyde and the FMNH2, which is provided by an FMN reductase such as LuxG. The LuxAB luciferase is a heterodimeric enzyme of almost 80kDa composed of α and β-subunits whose molecular weight is 42kDa and 39kDa. For the two subunits, the α subunit plays a major role which is responsible for the light-emitting reaction and the β-subunit is important for stabling the protein, although there is about 40% identity in the amino acid sequence between the α and β subunits.
Although some teams in previous years also designed coding sequences of LuxAB, but they did not provide enough information for users and did not prove that their parts could work well. However, we designed this part as a reporter used in our project by sequence optimization and the sequence we designed was finally synthesized by IDT as gBlock® Fragments. We also changed the RBS in this part, the natural RBS was replaced by iGEM BioBrick, BBa_B0034. With these designs, our designed part can work well.
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 530
- 1000INCOMPATIBLE WITH RFC[1000]Illegal SapI site found at 1049
Design Notes
This part was synthesized by IDT as gBlock® Fragments
Source
This part is from X.luminescens
References
[1] Thoden JB, Holden HM, Fisher AJ, Sinclair JF, Wesenberg G, Baldwin TO, Rayment I. Structure of the beta 2 homodimer of bacterial luciferase from Vibrio harveyi: X-ray analysis of a kinetic protein folding trap. Protein Sci. 1997 Jan;6(1):13-23.
[2] Lee CY, et al. The lux genes of the luminous bacterial symbiont, Photobacterium leiognathi, of the ponyfish. Nucleotide sequence, difference in gene organization, and high expression in mutant Escherichia coli. Eur J Biochem, 1991 Oct 1. PMID 1915359