Quantification, regulation and production of 5-aminolevulinic acid by green fluorescent protein in recombinant Escherichia coli

Tan, Shih‐I; You, Shao-Chun; Shih, I-Tai; Ng, I-Son

doi:10.1016/j.jbiosc.2019.10.005

Cited by 19 publications

(6 citation statements)

References 44 publications

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“…Actually, MM9 medium and artificial urine were at pH 7 and pH 5, respectively. According to the previous study, the sfGFP intensity was influenced by pH environment and reduced 55% at pH 5 . Thus, the poorer fluorescent signal in the artificial urine was attributed to the low pH value.…”

Section: Results and Discussionmentioning

confidence: 83%

CRISPRi-Mediated NIMPLY Logic Gate for Fine-Tuning the Whole-Cell Sensing toward Simple Urine Glucose Detection

Tan

2021

ACS Synth. Biol.

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Whole-cell biosensors have been regarded as a prominent alternative to chemical and physical biosensors due to their renewability, environmental friendliness, and biocompatibility. However, there is still a lack of noninvasive measurements of urine glucose, which plays a vital role in monitoring the risk of diabetes in the healthcare system, via whole-cell biosensors. In this study, we characterized a glucose-inducible promoter and further enhanced the sensing performance using three genetic effectors, which encompassed ribozyme regulator (RiboJ), clustered regularly interspaced short palindromic repeat interference (CRISPRi), and plasmid-based T7RNA polymerase (PDT7), to develop the noninvasive glucose biosensor by fluorescent signal. As a result, RiboJ increased dynamic range to 2989 au, but declined signal-to-noise (S/N) to 1.59, while CRISPRi-mediated NIMPLY gate intensified both dynamic range to 5720 au and S/N to 4.58. The use of single PDT7 orthogonal with T7 promoter in cells (i.e., P strain) achieved a 44 180 au of dynamic range with S/N at 3.08. By coupling the PDT7 and NIMPLY-mediated CRISPRi, we constructed an optimum PIGAS strain with the highest S/N value of 4.95. Finally, we adopted the synthetic bacteria into a microdevice to afford an integrative and portable system for daily urine glucose inspection, which would be an alternative approach for medical diagnosis in the future.

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Section: Results and Discussionmentioning

confidence: 83%

CRISPRi-Mediated NIMPLY Logic Gate for Fine-Tuning the Whole-Cell Sensing toward Simple Urine Glucose Detection

Tan

2021

ACS Synth. Biol.

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“…Kang et al firstly applied the rhtA gene in 5-ALA synthesis and successfully demonstrated its feasibility for further improvement of production (Kang et al, 2011a). Recently, overexpressing rhtA with a T7 system composed of orthogonal T7RNA polymerase and the T7 promoter increased 5-ALA production 40-fold in E. coli (Tan S. I. et al, 2019). Introduction of rhtA into the high-yield recombinant C. glutamicum AL-OI** increased production of 5-ALA by 28% over the control (Ko et al, 2019).…”

Section: Accelerated Transport Of Intracellular 5-aminolevulinic Acidmentioning

confidence: 99%

“…Although the concentration of 5-ALA is reflected in the fluorescence intensity, the precise concentration of 5-ALA still depends on Ehrlich's reagent. Recently, a fluorescence-based whole-cell system was developed for the simultaneous detection and production of 5-ALA in recombinant E. coli (Tan S. I. et al, 2019). In this method, a constitutive 5-ALA production strain with R. sphaeroides HemA (RshemA) and a fusion of super-folder-green fluorescent protein was established with an optimized promoter, medium, and fusion sites.…”

Section: Fluorescence Spectroscopymentioning

confidence: 99%

“…In this method, a constitutive 5-ALA production strain with R. sphaeroides HemA (RshemA) and a fusion of super-folder-green fluorescent protein was established with an optimized promoter, medium, and fusion sites. The de novo quantification of intracellular and extracellular 5-ALA by monitoring the fluorescence of sfGFP fused to RshemA was also explored using dual promoters and dual plasmids (Tan S. I. et al, 2019). Additionally, to increase the understanding of the regulatory mechanism of 5-ALA in higher organisms, matrix isopotential synchronous fluorescence spectrometry and the first derivative technique were combined for the direct determination of δ-ALA in urine samples (Ajmal et al, 2019).…”

Section: Fluorescence Spectroscopymentioning

confidence: 99%

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Natural 5-Aminolevulinic Acid: Sources, Biosynthesis, Detection and Applications

Jiang

Hong

Mao

et al. 2022

Front. Bioeng. Biotechnol.

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5-Aminolevulinic acid (5-ALA) is the key precursor for the biosynthesis of tetrapyrrole compounds, with wide applications in medicine, agriculture and other burgeoning fields. Because of its potential applications and disadvantages of chemical synthesis, alternative biotechnological methods have drawn increasing attention. In this review, the recent progress in biosynthetic pathways and regulatory mechanisms of 5-ALA synthesis in biological hosts are summarized. The research progress on 5-ALA biosynthesis via the C4/C5 pathway in microbial cells is emphasized, and the corresponding biotechnological design strategies are highlighted and discussed in detail. In addition, the detection methods and applications of 5-ALA are also reviewed. Finally, perspectives on potential strategies for improving the biosynthesis of 5-ALA and understanding the related mechanisms to further promote its industrial application are conceived and proposed.

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“…An extremely active enzyme can catalyze the reaction more rapidly as well as transform the substrates to products more efficiently. 5-Aminolevulinic acid synthase (ALAS), a key enzyme of the C4 pathway, is encoded by hem A gene from different organisms, such as Agrobacterium radiobacter (Fu et al 2010 ), Bradyrhizobium japonicum (Choi et al 1999 ; Jung et al 2004 ), Rhodobacter capsulatus (Kang et al 2004 ; Lou et al 2014 ; Yu et al 2020 ) , Paracoccus (Sato et al 1985 ), Rattus norvegicus (Nakakuki et al 1980 ), Rhodobacter sphaeroides (Kang et al 2011a , b ; Shih et al 2021 ; Tan et al 2020a ; Tran et al 2019 ), Rhodopseudomonas palustris (Ong et al 2013 ; Zhang et al 2013 ; Liu et al 2016 ), and Saccharomyces cerevisiae (Volland and Felix 1984 ). The ALAS from different organisms used for 5-ALA production via C4 pathway in the genetic E. coli strains are summarized in Table 1 .…”

Section: Production Of 5-ala Via C4 Pathway In Escherichia ...mentioning

confidence: 99%

Challenges and opportunities of bioprocessing 5-aminolevulinic acid using genetic and metabolic engineering: a critical review

Shih

et al. 2021

Bioresour. Bioprocess.

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Abstract5-Aminolevulinic acid (5-ALA), a non-proteinogenic five-carbon amino acid, has received intensive attentions in medicine due to its approval by the US Food and Drug Administration (FDA) for cancer diagnosis and treatment as photodynamic therapy. As chemical synthesis of 5-ALA performed low yield, complicated processes, and high cost, biosynthesis of 5-ALA via C4 (also called Shemin pathway) and C5 pathway related to heme biosynthesis in microorganism equipped more advantages. In C4 pathway, 5-ALA is derived from condensation of succinyl-CoA and glycine by 5-aminolevulic acid synthase (ALAS) with pyridoxal phosphate (PLP) as co-factor in one-step biotransformation. The C5 pathway involves three enzymes comprising glutamyl-tRNA synthetase (GltX), glutamyl-tRNA reductase (HemA), and glutamate-1-semialdehyde aminotransferase (HemL) from α-ketoglutarate in TCA cycle to 5-ALA and heme. In this review, we describe the recent results of 5-ALA production from different genes and microorganisms via genetic and metabolic engineering approaches. The regulation of different chassis is fine-tuned by applying synthetic biology and boosts 5-ALA production eventually. The purification process, challenges, and opportunities of 5-ALA for industrial applications are also summarized.

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Quantification, regulation and production of 5-aminolevulinic acid by green fluorescent protein in recombinant Escherichia coli

Cited by 19 publications

References 44 publications

CRISPRi-Mediated NIMPLY Logic Gate for Fine-Tuning the Whole-Cell Sensing toward Simple Urine Glucose Detection

CRISPRi-Mediated NIMPLY Logic Gate for Fine-Tuning the Whole-Cell Sensing toward Simple Urine Glucose Detection

Natural 5-Aminolevulinic Acid: Sources, Biosynthesis, Detection and Applications

Challenges and opportunities of bioprocessing 5-aminolevulinic acid using genetic and metabolic engineering: a critical review

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