Difference between revisions of "Part:BBa K2963009"

 
(8 intermediate revisions by the same user not shown)
Line 3: Line 3:
 
<partinfo>BBa_K2963009 short</partinfo>
 
<partinfo>BBa_K2963009 short</partinfo>
  
The gene cluster <i>capB*CA encode an multi-enzyme complex on cell membrane. This multi-enzyme complex is composed of capB*;capC and capA. capB catalyzes poly-γ-glutamic acid(γ-PGA) synthesis&#65294;And capC links CapB and CapA on the membrane. While CapA transports γ-PGA outside the cell.In our project, we use the this part to biosynthesize γ-PGA.
+
The gene cluster <i>capB*CA</i> encode an multi-enzyme complex which can polymorize D/L glutamic acid to synthesize γ-PGA.
 
+
 
===Usage and Biology===
 
===Usage and Biology===
The BCA genes from Bacillus sp., encoding a polyglutamate synthetase located on the cell membrane, is capable of polymerizing glutamic acid to form poly-γ-glutamic acid. In Bacillus licheniformis, BCA are called capBCA.This part is used for producing L-glutamate-rich γ-PGA.
+
The <i>BCA</i> genes from <i>Bacillus sp.</i>, encoding a polyglutamate synthetase located on the cell membrane, is capable of polymerizing glutamic acid to produce γ-PGA. In <i>Bacillus licheniformis</i>, <i>BCA</i> are called <i>capBCA</i>. We used the mutant <i>B* </i>gene of <i>B</i> gene. This part is used for producing L-glutamate-rich γ-PGA.
  
 
===Characterization===
 
===Characterization===
We used NMR to detect γ-PGA and HPLC to analyze L- glutamate ratio of γ-PGA. The results showed we have successfully produced L-glutamate-rich γ-PGA.
+
We used NMR to detect γ-PGA and HPLC to analyze L- glutamate ratio of γ-PGA we have produce. The results as follow show that we have successfully produced L-glutamate-rich γ-PGA.
  
 
[[image:NMR.png|400px]]
 
[[image:NMR.png|400px]]
  
By the NMR detection of the fermentation product, the specific hydrogen peaks a, b, and c on the γ-amide bond on the γ-polyglutamic acid could be detected at the corresponding time points. It was indicated that the synthetase capB*CA genes of Bacillus licheniformis heterologously expressed in Corynebacterium glutamicum, and the target product γ-polyglutamic acid was successfully produced.
+
By the NMR detection of the fermentation product, the specific hydrogen peaks a, b, and c on the γ-amide bond on the γ-polyglutamic acid could be detected at the corresponding time points. It is indicated that the <i>capB*CA</i> genes from <i>Bacillus licheniformis</i> heterologously express in <i>Corynebacterium glutamicum</i>, and the target product γ-PGA is successfully produced.
  
[[image:HPLC.png|400px]]
 
  
 
We used HPLC to detect the L-glutamate monomer ratio of the γ-PGA we have produced. The results show that the L-glutamic acid monomer ratio reaches more than 90%. This part is working and we have produced L-glutamate-rich γ-PGA.  
 
We used HPLC to detect the L-glutamate monomer ratio of the γ-PGA we have produced. The results show that the L-glutamic acid monomer ratio reaches more than 90%. This part is working and we have produced L-glutamate-rich γ-PGA.  
  
 +
[[image:HPLC.png|400px]]
 +
 +
===References===
 +
1. Xu P, Vansiri A, Bhan N, et al. ePathBrick: a synthetic biology platform for engineering metabolic pathways in E. coli[J]. ACS Synthetic Biology, 2012, 1(7): 256-266.
 +
 +
2. Peng Yingyun. Study on the production, synthesis mechanism and antifreeze of γ-polyglutamic acid. Diss. Jiangnan University, 2015.
 +
 +
3. Sung M H, Park C, Kim C J, et al. Natural and edible biopolymer poly-gamma-glutamic acid: synthesis, production, and applications [J]. Chemical Record, 2005, 5(6): 352-366.
  
 
<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here

Latest revision as of 14:54, 19 October 2019


capB*CA - encoding poly-γ-glutamic acid synthetase

The gene cluster capB*CA encode an multi-enzyme complex which can polymorize D/L glutamic acid to synthesize γ-PGA.

Usage and Biology

The BCA genes from Bacillus sp., encoding a polyglutamate synthetase located on the cell membrane, is capable of polymerizing glutamic acid to produce γ-PGA. In Bacillus licheniformis, BCA are called capBCA. We used the mutant B* gene of B gene. This part is used for producing L-glutamate-rich γ-PGA.

Characterization

We used NMR to detect γ-PGA and HPLC to analyze L- glutamate ratio of γ-PGA we have produce. The results as follow show that we have successfully produced L-glutamate-rich γ-PGA.

NMR.png

By the NMR detection of the fermentation product, the specific hydrogen peaks a, b, and c on the γ-amide bond on the γ-polyglutamic acid could be detected at the corresponding time points. It is indicated that the capB*CA genes from Bacillus licheniformis heterologously express in Corynebacterium glutamicum, and the target product γ-PGA is successfully produced.


We used HPLC to detect the L-glutamate monomer ratio of the γ-PGA we have produced. The results show that the L-glutamic acid monomer ratio reaches more than 90%. This part is working and we have produced L-glutamate-rich γ-PGA.

HPLC.png

References

1. Xu P, Vansiri A, Bhan N, et al. ePathBrick: a synthetic biology platform for engineering metabolic pathways in E. coli[J]. ACS Synthetic Biology, 2012, 1(7): 256-266.

2. Peng Yingyun. Study on the production, synthesis mechanism and antifreeze of γ-polyglutamic acid. Diss. Jiangnan University, 2015.

3. Sung M H, Park C, Kim C J, et al. Natural and edible biopolymer poly-gamma-glutamic acid: synthesis, production, and applications [J]. Chemical Record, 2005, 5(6): 352-366.