Part:BBa_K4275036

pLAC4-KmarxMFα-NpaBGS-t-tTDH1

The composite part contains a pLac4 promoter[1], Kluyveromyces marxianus mating factor alpha[2], beta-glucosidase NpaBGS fused with type I dockerin[3] and tTDH1 terminator[4]. NpaBGS is a cellulase active on oligosaccharides(short polysaccharides) and cellobiose(disaccharide) that cleaves glycosidic bonds. The Kluyveromyces marxianus mating factor alpha is fused in order to enable secretion of NpaBGS by host K.marxianus.

Usage and Biology

Kluyveromyces marxianus mating factor alpha fused with NpaBGS would lead the polypeptide of NpaBGS-t through ER and Golgi apparatus and be secreted. NpaBGS is a β-glucosidase containing two glycosyl hydrolase Family 3 domains at N and C terminals .It was discovered from the buffalo rumen fungus Neocallimastix patriciarum W5. It’s activity could be enhanced at 50°C by adding Mg2+ or Mn2+ ions.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal EcoRI site found at 331
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal EcoRI site found at 331
  Illegal NheI site found at 89
  Illegal NheI site found at 1783
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal EcoRI site found at 331
  Illegal XhoI site found at 1252
  Illegal XhoI site found at 2858
  Illegal XhoI site found at 3937
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal EcoRI site found at 331
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal EcoRI site found at 331
  Illegal NgoMIV site found at 2355
  Illegal AgeI site found at 1369
  Illegal AgeI site found at 1387
  
• 1000
  COMPATIBLE WITH RFC[1000]

References

1. Rajkumar, Arun S. et al. "Biological Parts For Kluyveromyces Marxianus Synthetic Biology". Frontiers In Bioengineering And Biotechnology, vol 7, 2019. Frontiers Media SA, https://doi.org/10.3389/fbioe.2019.00097.
2.“Mating Factor Alpha-1 [Kluyveromyces Marxianus] - Protein - NCBI.” National Center for Biotechnology Information, U.S. National Library of Medicine, https://www.ncbi.nlm.nih.gov/protein/QGN17207.1
3. Chen, Hsin-Liang et al. "A Highly Efficient Β-Glucosidase From The Buffalo Rumen Fungus Neocallimastix Patriciarum W5". Biotechnology For Biofuels, vol 5, no. 1, 2012. Springer Science And Business Media LLC, https://doi.org/10.1186/1754-6834-5-24.
4. "Part:Bba K2753052 - Parts.Igem.Org". Parts.Igem.Org, 2022, https://parts.igem.org/Part:BBa_K2753052.