Team:BGIC-Global/Parts

PARTS

Parts
BBa_K2728001 - pFrmR

Part type: promotor

Description:
This promoter is an engineered formaldehyde-inducible promoter. Escherichia coli has a native formaldehyde-inducible promoter, pfrm, which is found upstream of the frmRAB formaldehyde detoxification operon. FrmR, the first product of the operon, is a member of the DUF156 family of DNA-binding transcriptional regulators. It binds the frmRAB promoter region and is negatively allosterically modulated by formaldehyde. FrmR is specific to formaldehyde, responding to acetaldehyde, methylglyoxal, and glyoxal to far lesser degrees and not at all to a range of other aldehydes and alcohols tested. The genes frmA and frmB encode a formaldehyde dehydrogenase and S-formylglutathione hydrolase, respectively, and are responsible for detoxifying formaldehyde to formic acid in a glutathione-dependent pathway. The negative-feedback regulation of the frmRAB operon is similar to that of many other prokaryotic operons, whereby the transcription factor represses its own transcription.

Feature:
  • 1. It’s a formaldehyde-inducible promoter from E.coli.
  • 2. It’s an engineered promoter. It retains formaldehyde responsiveness, with 2-fold higher GFP expression in response to 100 μM formaldehyde than the native pfrm. Application of this promoter with higher basal and induced expression levels before methanol assimilation genes achieves higher biomass titers than the native E. coli pfrm.
    • BBa_K2728002 - EGFP

    Part type: coding

    Description:
    BBa_K2728002 is a basic (constitutively fluorescent) green fluorescent protein published in 1996, derived from Aequorea victoria. It is reported to be a rapidly-maturing weak dimer with moderate acid sensitivity.
    BBa_K2728003 - Nanoluc

    Part type: coding

    Description:
    BBa_K2728003 is a small enzyme (19.1kDa) engineered for optimal performance as a luminescent reporter. The enzyme is about 100-fold brighter than either firefly (Photinus pyralis) or Renilla reniformis luciferase using a novel substrate, furimazine, to produce high intensity, glow-type luminescence. The luminescent reaction is ATP-independent and designed to suppress background luminescence for maximal assay sensitivity.

    Sequence and Features:
  • BBa_K2728003 possesses a number of physical properties that make it an excellent reporter protein:
  • 1. very small, monomeric enzyme (171 amino acids; 513bp)
  • 2. high thermal stability (Tm = 60°C)
  • 3. active over a broad pH range (pH 6–8)
  • 4. no post-translational modifications or disulfide bonds
  • 5. uniform distribution in cells
  • 6. emission spectrum well suited for bioluminescence resonance energy transfer (BRET; λmax = 465nM)
    • BBa_K2728004 - GFA

    Part type: coding

    Basic Description:
    BBa_K2728004 is an enzyme from Paracoccus denitrificans that accelerates this spontaneous condensation reaction, which catalyze the conversion of formaldehyde and glutathione was purified and named glutathione-dependent formaldehyde-activating enzyme (Gfa). BBa_K2728004 is located directly upstream of the gene for glutathione-dependent formaldehyde dehydrogenase, which catalyzes the subsequent oxidation of S-hydroxymethylglutathione. The glutathione-dependent formaldehyde conversion to formate starts with the adduct formation, formaldehyde reacts with the SH group of glutathione producing S-hydroxymethylglutathione .
    *We guarantee all parts are tested and proved by us before the parts submission that iGEMers can use them directly.
    2018@BGIC-Global