Part:BBa_K4229018

RBS-spyCatcherXiaI-RBS-TnaB

This composite part was used to for the coexpression of two target genes of the indigo/indirubin pathways. The first one is XiaI fused with the SpyCatcher (BBa_K4229015) at it N-terminus and the second is the L-tryptophan transport protein TnaB (BBa_K4229003).

These two genes build together with the BBa_K4229016 all enzymes of the indigo/indirubin pathway.

Figure 1: Schematic representation of the indigo/indirubin pathway. L-tryptophan is imported by the membrane protein TnaB (low affinity tryptophan permease). L-tryptophan is cleaved into indole, NH4+ and pyruvate by the tryptophanase TnaA. The reaction continues by the hydroxylation of indole through XiaI. To enhance the effectivity of this enzyme, the NAD(P)H-flavin reductase provides XiaI with FADH2 by adding hydrogen to FAD. Finally, indole is transformed to either 3-Hydroxyindole or 2-Hydroxyindole. These two substances spontaneously react to 3-Oxindole and 2-Oxindole through the secession of hydrogen from the OH-group. Through spontaneous dimerization indigo and indirubin are formed. Graphic adapted from [1].

Usage and Biology

The production was tested under the T7 promotor and LacI operator in those two Biobricks: BBa_K4229033 and BBa_K4229034

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 893
  Illegal AgeI site found at 783
  
• 1000
  COMPATIBLE WITH RFC[1000]