Parts

Summary

Our project aims to develop a modified E. coli Nissle 1917 (EcN1917) that can persistently produce arginine in the tumor microenvironment (TME). Native EcN1917 has pathways for arginine synthesis but they are inhibited by the feedback of arginine in the environment. Therefore, to reach our goals, we constructed EcN1917 to maximize arginine production by avoiding these feedback inhibitions. The parts we constructed this year could be divided in two systems, as followed.

argR Knock-out system

Our project aims to develop a modified E. coli Firstly, we knocked out the argR gene which encodes ArgR protein. ArgR represses the expression of enzyme genes important for arginine biosynthesis in response to the arginine level in the cell. To achieve this goal, we successfully constructed 6 parts: BBa_K4410000-BBa_K4410003, BBa_K4410005, and BBa_K4410006.

Arginine production biobrick

We also added the argJ gene derived from Corynebacterium to remove the arginine inhibition on NAGS enzyme encoded by argA gene. argJ encodes bifunctional glutamate N-acetyltransferase/amino-acid acetyltransferase, and can achieve the same function of acetylation of glutamate as NAGS at high concentration of arginine. To achieve this goal, we successfully constructed 3 parts: BBa_K4410004, BBa_K4410007, and BBa_K4410008.

No.	Name	Type	Description	Designer	Length
1	BBa_K4410000	Basic	Fragment H1 from E. coli Nissle 1917	Ruyi Shi	1000bp
2	BBa_K4410001	Basic	Fragment H2 from E. coli Nissle 1917	Ruyi Shi	1000bp
3	BBa_K4410002	Basic	argR	Ruyi Shi	472bp
4	BBa_K4410003	Basic	KanR	Ruyi Shi	795bp
5	BBa_K4410004	Basic	argJ	Ruyi Shi	1167bp
6	BBa_K4410005	Composite	H1-argR-H2	Ruyi Shi	2472bp
7	BBa_K4410006	Composite	H1-KanR-H2	Ruyi Shi	2795bp
8	BBa_K4410007	Device	J23100-argJ	Ruyi Shi	1334bp
9	BBa_K4410008	Basic	Terminator	Ruyi Shi	87bp