Part:BBa_K2242018

T7+lacO+Mtr

Mtr CAB is a protein complex located on the outer membrane of Shewanella originally, transferring electrons from the cityplasm to the outside of the bacteria. However, according to the lately research, we found that this Mtr CAB protein complex can also transport electrons into the cytoplasm from the electrode or other electron donors from the outside. This is the role Mtr CAB plays in our project. We introduce this protein complex into our engineered E.coli to transport electrons into the cytoplasm. Mtr CAB consists of three proteins, Mtr A, Mtr B, Mtr C. Mtr B is anchored onto the outer membrane. With its ß barrel conformation, it can help to locate the Mtr A and Mtr C and increase the whole complex’s stability. Mtr A and Mtr C are the two protein that can actually transfer electrons with the heme attached into these two protein at the right position. MtrA is a 32-kD periplasmic decaheme cytochrome c, and MtrC is a 69-kD cell-surface-exposed. Electrons are collected by Mtr C then it will shuttle through this electron tunnel and go to Mtr A, then these electrons will be transferred to CymA on the inner membrane and final get to the NADH dehydrogenase through the electron transfer chain. With this protein complex, we can utilize the electrons from the electrode or some chemical compound outside of the bacteria can turn them into NADH when they get into the cytoplasm of our engineered bacteria, increasing the NADH’s concentration inside of the cytoplasm, which means their the reduction power ——the ability to synthesize, will be pump up. As we have mentioned, it’s the heme attached to the Mtr A and Mtr C that enable them to shuttle electrons. So these two protein belong to a protein family called cytochrome c. We use T7 promoter and Lac operator to control Mtr CAB’s expression. Both of this two units are induced by IPTG. With this dual switch, we can reduce the leak of the gene expression as much as possible.

Sequence and Features

Assembly Compatibility:

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

  Illegal XhoI site found at 5267
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 309