Design and Application of an Artificial Hybrid PromoterPluxI-lacOin Genetic Circuit to Achieve Lower Basal Expression Level

Huang, Shiyang; Song, Yi-Hang; Zhuang, Xinbo; Gao, Ze-Yue; Wang, Ke; Peng, Yajuan; Fang, Baishan

doi:10.1007/s12010-019-03153-4

Cited by 2 publications

(3 citation statements)

References 28 publications

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“…55 In another study, by combining the feature of the lac operon and the regulation of the QS, a hybrid promoter suitable for the efficient screening of quorum sensing-disrupting enzymes was created. 56 Establishing HTS methods for the identification of better producer strains The capability of synthesizing target compounds varies from cell to cell in synthetic biology studies. The extent to which the desired strain is obtained from a mutant library created through random mutagenesis is primarily affected by the availability of HTS.…”

Section: Application Of Engineered Tfs Constructing Inducible Expressmentioning

confidence: 99%

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Transcriptional factor engineering in microbes for industrial biotechnology

Wang

Liang

Xing

et al. 2020

J of Chemical Tech & Biotech

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Transcription can be regulated by controlling the provision of transcriptional factors (TFs), because TFs determine the transcription process by specifying the binding of RNA polymerase holoenzyme on promoters. Manipulations, such as replacing endogenous TFs with mutants obtained through directed evolution (or rational design), and introducing compatible heterogeneous TFs, are effective ways to regulate transcription. Designing artificial TFs with desired properties by following the principles of protein design and reconstructing ancestral TFs with the information gained from a perspective of evolution by multiple sequence alignments of the related TFs, are feasible alternative options. The engineered TFs can be used to create inducible protein expression systems that respond to a specific stimulus. Moreover, the engineered TFs could lead to a transcriptional profile different from that of the wild-type strains. Accordingly, these engineered TFs could be utilized to construct strains resistant to adverse conditions, since the emergence of resistance generally requires global transcriptional changes at the transcriptomic scale. Meanwhile, these engineered TFs can also be employed in the construction of sophisticatedly designed genetic circuits for diverse purposes. In this paper, we reviewed the advances in the above-mentioned aspects.

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Section: Application Of Engineered Tfs Constructing Inducible Expressmentioning

confidence: 99%

“…In these two designed gene circuits, induction was achieved in a dose‐dependent manner with 0.05 to 10 μM aspirin 55 . In another study, by combining the feature of the lac operon and the regulation of the QS, a hybrid promoter suitable for the efficient screening of quorum sensing‐disrupting enzymes was created 56 …”

Section: Application Of Engineered Tfsmentioning

confidence: 99%

Transcriptional factor engineering in microbes for industrial biotechnology

Wang

Liang

Xing

et al. 2020

J of Chemical Tech & Biotech

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“…Based on semi‐rational approaches, LacI has been engineered to alter its effector specificities, responding to chemicals other than its natural effectors, such as fucose, gentiobiose, lactitol, sucralose and lactulose (Taylor et al., 2016 ; Wu et al., 2017 ). Instead, to decrease basal expression, LacI Q1 promoter or its derivatives (Calos, 1978 ; Calos & Miller, 1981 ; Glascock & Weickert, 1998 ), and a hybrid promoter of lacI (Huang et al., 2020 ), were widely employed in recombinant expression. LacI Q1 promoter leads to a 50‐ to 100‐fold increased expression of lac repressor, greatly repressing the leaky expression because the degree of repression is supposedly dependent on the ratio of intracellular LacI molecules to the lacO operator sites (Calos & Miller, 1981 ; Glascock & Weickert, 1998 ; Raibaud & Schwartz, 1984 ).…”

Section: Introductionmentioning

confidence: 99%

Engineering and application of LacI mutants with stringent expressions

Wu,

Liang,

et al. 2024

Microbial Biotechnology

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Optimal transcriptional regulatory circuits are expected to exhibit stringent control, maintaining silence in the absence of inducers while exhibiting a broad induction dynamic range upon the addition of effectors. In the Plac/LacI pair, the promoter of the lac operon in Escherichia coli is characterized by its leakiness, attributed to the moderate affinity of LacI for its operator target. In response to this limitation, the LacI regulatory protein underwent engineering to enhance its regulatory properties. The M7 mutant, carrying I79T and N246S mutations, resulted in the lac promoter displaying approximately 95% less leaky expression and a broader induction dynamic range compared to the wild‐type LacI. An in‐depth analysis of each mutation revealed distinct regulatory profiles. In contrast to the wild‐type LacI, the M7 mutant exhibited a tighter binding to the operator sequence, as evidenced by surface plasmon resonance studies. Leveraging the capabilities of the M7 mutant, a high‐value sugar biosensor was constructed. This biosensor facilitated the selection of mutant galactosidases with approximately a seven‐fold improvement in specific activity for transgalactosylation. Consequently, this advancement enabled enhanced biosynthesis of galacto‐oligosaccharides (GOS).

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Design and Application of an Artificial Hybrid PromoterPluxI-lacOin Genetic Circuit to Achieve Lower Basal Expression Level

Cited by 2 publications

References 28 publications

Transcriptional factor engineering in microbes for industrial biotechnology

Transcriptional factor engineering in microbes for industrial biotechnology

Engineering and application of LacI mutants with stringent expressions

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