Part:BBa_K4229013

TnaA with N-terminal SnoopCatcher

TnaA belongs to the family of tryptophanase (tna) operon leader peptide. Tryptophanase catalyses the degradation of L-tryptophan to indole, pyruvate and ammonia, enabling the bacteria to utilise tryptophan as a source of carbon, nitrogen and energy. The tna operon of E. coli contains two major structural genes, tnaA and tnaB.

SnoopCatcher (BBa_K4229009) is fused via a small linker N-terminal TnaA. With that, we made it able to be recruited into the microcompartment (wiffelball).

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 [2].

References
[1]. A transcriptional pause synchronizes translation with transcription in the tryptophanase operon leader region. Gong F, Yanofsky C. J. Bacteriol. 185, 6472-6, (2003). View articlePMID: 14563884
[2] H. Yin et al., “Efficient Bioproduction of Indigo and Indirubin by Optimizing a Novel Terpenoid Cyclase XiaI in Escherichia coli,” ACS Omega, vol. 6, no. 31, pp. 20569–20576, 2021, doi: 10.1021/acsomega.1c02679.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal PstI site found at 977
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal PstI site found at 977
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal PstI site found at 977
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal PstI site found at 977
  Illegal AgeI site found at 142
  Illegal AgeI site found at 228
  Illegal AgeI site found at 1600
  
• 1000
  COMPATIBLE WITH RFC[1000]