Part:BBa_K3102044:Design

Electron transfer improvement (ACS, ZWF, GND, PGL)

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal PstI site found at 2933
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal PstI site found at 2933
  Illegal NotI site found at 2814
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BamHI site found at 3490
  Illegal XhoI site found at 408
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal PstI site found at 2933
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal PstI site found at 2933
  Illegal NgoMIV site found at 2206
  Illegal NgoMIV site found at 2923
  Illegal NgoMIV site found at 5493
  Illegal AgeI site found at 837
  Illegal AgeI site found at 4245
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal SapI.rc site found at 3975
  

Design Notes

In the ACS, we mutated a Cytosine into Thymine at the 1563bp position (C1563T) because of the Esp3I enzyme that we used for our Golden Gate Assembly (GGA) construct. (e.g BBa_K3102021)

In the ZWF, we mutated a Thymine into Cysteine at the 844bp position (T844C) because of the PstI enzyme (e.g. BBa_K3102008)

Source

RibA : E.coli, sequence found through Biocyc https://biocyc.org/gene?orgid=ECOLI&id=EG11331

RibB: E.coli, sequence found through Biocyc https://biocyc.org/gene?orgid=ECOLI&id=EG10465

RibD: E.coli, sequence found through Biocyc https://biocyc.org/gene?orgid=ECOLI&id=EG11321

RibE: E.coli, sequence found through Biocyc https://biocyc.org/gene?orgid=ECOLI&id=EG11322

RibC: E.coli, sequence found through Biocyc https://biocyc.org/gene?orgid=ECOLI&id=EG11406

RibF: E.coli, sequence found through Biocyc https://biocyc.org/gene?orgid=ECOLI&id=EG11079

AWS: E.coli, sequence found through Biocyc https://biocyc.org/gene?orgid=ECOLI&id=EG11448

ZWF: Corynebacterium glutamicum, sequence found through Ebi https://www.ebi.ac.uk/ena/data/view/BAB98969

GND: Corynebacterium glutamicum, sequence found through GenBank https://biocyc.org/gene?orgid=CORYNE&id=G18NG-11035-MONOMER#tab=TU

GLD: E.coli, sequence found through GenBank https://www.ncbi.nlm.nih.gov/nuccore/NC_000913.3?from=798586&to=799581&report=fasta

Promoter (BBa_I719005), RBS (BBa_B0034) and Terminator (BBa_B0015) are from the iGEM Registry.

References

Bacher A. et al., "Riboflavin synthases of Bacillus subtilis. Purification and properties." Biol. Chem. 255:632-637(1980)

Brown TD. et al., The enzymic interconversion of acetate and acetyl-coenzyme A in Escherichia coli, J Gen Microbiol. 1977 Oct;102(2):327-36.

Chen Y.Y. et al., "Conformational changes associated with cofactor/substrate binding of 6-phosphogluconate dehydrogenase from Escherichia coli and Klebsiella pneumoniae: Implications for enzyme mechanism.", J. Struct. Biol. 169:25-35(2010)

Eberhardt S.M.R. et al., J"Cloning, sequencing, mapping and hyperexpression of the ribC gene coding for riboflavin synthase of Escherichia coli." . Biochem. 242:712-719(1996)

Kumari S. et al., Cloning, characterization, and functional expression of acs, the gene which encodes acetyl coenzyme A synthetase in Escherichia coli, J Bacteriol. 1995 May;177(10):2878-86.

Lin Z, et al., "Metabolic engineering of Escherichia coli for the production of riboflavin". Microb Cell Fact. 2014;13(1):1–12.

Thomason L.C., Court D.L. et al., "Identification of the Escherichia coli K-12 ybhE gene as pgl, encoding 6-phosphogluconolactonase." Bacteriol. 186:8248-8253(2004)

Zhao J. et al., "Effect of zwf gene knockout on the metabolism of Escherichia coli grown on glucose or acetate." Metab. Eng. 6:164-174(2004)

Zhenquan Lin et al., “Metabolic engineering of Escherichia coli for the production of riboflavin," Microbial Cell Factories, 2014, 13:104.