Part:BBa_K3596030
BIOT: PpdD Signal+Pilin Gene+ TypeIV pilin assembly system
BIOT is a basic design for the expression of Geobacter sulfurreducens' pilin-based electrically conductive protein nanowires (e-PNs)[1]. The system includes a tac promoter which promotes the pilin gene(pilA) to express protein monomers; the monomers are later assembled into protein nanowires by the TypeIV pilin assembly system, which is constructed from the gene cluster at the downstream of the pilin gene(figure 1). The e-PNs can generate electric potential with the presence of water molecules in the air, which can be further modified for other purposes.
Characterization
The e-PN can be successfully expressed by express this part(figure 1)
.
Figure 1: The Genetic circuit of BIOT
Then we followed the protocol to purify the target protein(figure 2), gaining protein with a concentration ranging from 200 to 400 μg/ml(figure 3).
Figure 2: The procedures for BIOT nanowires purification.
Figure 3: The Protein Concentration of BIOT purification.
Western Blot was used to confirm the product protein as our targeting one, and the result showed it clearly (figure 4).
Figure 6: The Structure of e-PN monomer protein(A) and Western blot to verify the expression of BIOT monomers;(B).
We made a protein film following the instructions from the paper we researched(figure 5).[2]
Figure 5: BIOT membrane on the transparent membrane.
Then we successfully built up the basic module of BIOT (figure 6) using this protein film.The gold positive electrode at the bottom is larger so that the e-PNs can be positioned on it. And the tiny circular carbon piece works as a negative electrode on the top.
Figure 6: BIOT Assembly and Measurement
We eventually succeeded in producing stable voltage at about 0.35V with a current of 500nA or so (under 60% relative humidity)(figure 7 A,B).Additionally, to find the optimum relative humidity for the battery to function, we did the measurement with different relative humidities. The optimum RH was roughly at 40-50%, where the voltage rises to about 0.42V (figure 7C).
Figure 7: The Key parameters of BIOT basic module. A, The voltage of BIOT; B, the current of BIOT; C, Voltage of BIOT under different relative humidities.
References
[1] Ueki, Toshiyuki, et al. An Escherichia Coli Chassis for Production of Electrically Conductive Protein Nanowires. ACS Synthetic Biology, vol.9, no.3, 3 Mar.2020, pp.647–654.
[2] Liu, Xiaomeng, et al.. Power Generation from Ambient Humidity Using Protein Nanowires. Nature, vol.578, no.7796, Feb.2020, pp.550–554.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 2393
Illegal BglII site found at 2857
Illegal BglII site found at 4658
Illegal BamHI site found at 1073
Illegal BamHI site found at 3182 - 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 1300
Illegal NgoMIV site found at 6145
Illegal AgeI site found at 1146
Illegal AgeI site found at 1439
Illegal AgeI site found at 2002
Illegal AgeI site found at 4626
Illegal AgeI site found at 6914 - 1000INCOMPATIBLE WITH RFC[1000]Illegal SapI site found at 7920
Illegal SapI.rc site found at 4262