PMID- 36087056
OWN - NLM
STAT- MEDLINE
DCOM- 20230330
LR  - 20230330
IS  - 2161-5063 (Electronic)
IS  - 2161-5063 (Linking)
VI  - 11
IP  - 10
DP  - 2022 Oct 21
TI  - Reprogramming T7RNA Polymerase in Escherichia coli Nissle 1917 under Specific Lac 
      Operon for Efficient p-Coumaric Acid Production.
PG  - 3471-3481
LID - 10.1021/acssynbio.2c00363 [doi]
AB  - Lac operon is the standard regulator used to control the orthogonality of T7RNA 
      polymerase (T7RNAP) and T7 promoter inEscherichia coli BL21(DE3) strain for 
      protein expression. However,E. coliNissle 1917 (EcN), the unique probiotic 
      strain, has seldom been precisely adapted to the T7 system. Herein, we applied 
      bioinformatics analysis on Lac operon from different strains, and it was observed 
      that a weak promoter for LacI repressor existed in EcN. Furthermore, X-gal assay 
      revealed a strong expression of lacZ in EcN. We demonstrated that Lac operon 
      significantly affected the protein expression in the two T7-derived EcN, in which 
      T7RNAP was integrated at lambda (ET7L) and HK022 (ET7H), respectively. Different 
      combinations of replication origin, chaperonin GroELS, inducer, and medium were 
      explored to fine-tune the best strain with tyrosine ammonia-lyase (TAL) for 
      p-coumaric acid (pCA) production, which is one of the essential bioactive 
      compounds for human health. Finally, the highest pCA conversion of 78.8% was 
      achieved using RRtL (plasmid form) under the optimum condition, and a 51.5% 
      conversion was obtained with L::Rt strain which has integrated T7-RtTAL at HK022 
      of ET7L in the simulated gut environment. The appropriate reprogramming of T7RNAP 
      expedites EcN as an effective and promising cell factory for live bacterial 
      therapeutics in the future.
FAU - Effendi, Sefli Sri Wahyu
AU  - Effendi SSW
AD  - Department of Chemical Engineering, National Cheng Kung University, Tainan 70101, 
      Taiwan.
FAU - Ng, I-Son
AU  - Ng IS
AUID- ORCID: 0000-0003-1659-5814
AD  - Department of Chemical Engineering, National Cheng Kung University, Tainan 70101, 
      Taiwan.
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
DEP - 20220910
PL  - United States
TA  - ACS Synth Biol
JT  - ACS synthetic biology
JID - 101575075
RN  - EC 2.7.7.6 (DNA-Directed RNA Polymerases)
RN  - IBS9D1EU3J (p-coumaric acid)
SB  - IM
MH  - Humans
MH  - *DNA-Directed RNA Polymerases/genetics/metabolism
MH  - *Escherichia coli/genetics/metabolism
MH  - Lac Operon
MH  - Operon/genetics
OTO - NOTNLM
OT  - E. coli Nissle
OT  - T7RNA polymerase
OT  - chromosomal integration
OT  - lac operon
OT  - loci effect
OT  - p-coumaric acid
EDAT- 2022/09/11 06:00
MHDA- 2022/10/25 06:00
CRDT- 2022/09/10 09:03
PHST- 2022/09/11 06:00 [pubmed]
PHST- 2022/10/25 06:00 [medline]
PHST- 2022/09/10 09:03 [entrez]
AID - 10.1021/acssynbio.2c00363 [doi]
PST - ppublish
SO  - ACS Synth Biol. 2022 Oct 21;11(10):3471-3481. doi: 10.1021/acssynbio.2c00363. 
      Epub 2022 Sep 10.