Part:BBa_K4613014

T3-ADH3

T3(BBa_K4613011) and C3(BBa_K4613012) could form protein complexes by elastin-like polypeptides(ELPs) monomers containing SpyTags and SpyCatchers. If you want to learn about the detailed introduction of T3 and C3, you can click the link below. https://parts.igem.org/Part:BBa_K4164011 https://parts.igem.org/Part:BBa_K4164012 Different functional proteins can be incorporated into the polymeric scaffolds in a flexible manner due to its programmability. In this part, NAU-CHINA 2023 incorporated ADH3, which is the most efficient OTA-detoxifying enzyme reported thus far and can hydrolyze OTA to nontoxic ochratoxin α(OTα) and L-β-phenylalanine(Phe). Moreover, its soluble protein expression of ADH3 in Escherichia coli has been realized. We fused ADH3 into T3 to immobilize the enzyme and increase the stability and sustainable production of ADH3.

Fig. 1 SDS-PAGE analysis of protein expression trials in E. coli BL21 (DE3) cultured in LB medium for 12 hours using pET29a(+)-T3-ADH3. The temperature was 20 °C. Lane M：protein marker. Lane 1: induced total protein. Lane 2: precipitate. Lane 3: supernatant.

Reference

1. Dai Z, Yang X, Wu F, et al.Living fabrication of functional semi-interpenetrating polymeric materials[J].Nat Commun,2021, 12 (1): 3422.
2. Zakeri B, Fierer J O, Celik E, et al.Peptide tag forming a rapid covalent bond to a protein, through engineering a bacterial adhesin[J].Proc Natl Acad Sci U S A,2012, 109 (12): E690-7.
3. Reddington S C, Howarth M.Secrets of a covalent interaction for biomaterials and biotechnology: SpyTag and SpyCatcher[J].Curr Opin Chem Biol,2015, 29: 94-9.
4. Dai L, Niu D, Huang J W, et al.Cryo-EM structure and rational engineering of a superefficient ochratoxin A-detoxifying amidohydrolase[J].J Hazard Mater,2023, 458: 131836.

Sequence and Features

Assembly Compatibility:

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

  Illegal BamHI site found at 940
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 1675
  Illegal AgeI site found at 1363
  Illegal AgeI site found at 1525
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal SapI site found at 8