Difference between revisions of "Part:BBa K5034213"
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| − | + | ===Basic Description=== | |
This basic part encodes the PPK1 gene which is initially from Citrobacter freundii and we performed codon optimization on, is expressed in the PYYDT plasmid. This basic part is designed to facilitate the reversible conversion between inorganic polyphosphate (PolyP) and inorganic phosphate (Pi). The PPK1 enzyme is known for its ability to synthesize PolyP from ATP and to degrade PolyP back to Pi, with a preference for the synthetic reaction, making it a versatile tool for managing phosphate metabolism in engineered systems. | This basic part encodes the PPK1 gene which is initially from Citrobacter freundii and we performed codon optimization on, is expressed in the PYYDT plasmid. This basic part is designed to facilitate the reversible conversion between inorganic polyphosphate (PolyP) and inorganic phosphate (Pi). The PPK1 enzyme is known for its ability to synthesize PolyP from ATP and to degrade PolyP back to Pi, with a preference for the synthetic reaction, making it a versatile tool for managing phosphate metabolism in engineered systems. | ||
| − | + | In a sentence, it can reversibly convert Poly p and Pi. This reversible process favors the generation of Poly P.For the first time, we expressed this element in a strain of Shewanella and conducted codon optimization based on Shewanella. | |
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Figure 1: Basic function of PPK1 | Figure 1: Basic function of PPK1 | ||
| − | + | ===Construct features(only coding sequence included in basic part)=== | |
Promoter: Constitutive promoter for continuous expression. We use tac promoter in our experiment. | Promoter: Constitutive promoter for continuous expression. We use tac promoter in our experiment. | ||
PPK1 Coding Sequence: Encodes the polyphosphate kinase 1 enzyme. | PPK1 Coding Sequence: Encodes the polyphosphate kinase 1 enzyme. | ||
Terminator: Efficient transcription terminator to ensure proper mRNA processing. We use T7Te terminator in our experiment. | Terminator: Efficient transcription terminator to ensure proper mRNA processing. We use T7Te terminator in our experiment. | ||
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Figure 2: Basic construction of PPK1 plasmid | Figure 2: Basic construction of PPK1 plasmid | ||
| − | + | ===Origin (Organism)=== | |
The PPK1 gene was sourced from Citrobacter freundii. The PYYDT plasmid backbone is a standard vector used for gene expression in synthetic biology applications. | The PPK1 gene was sourced from Citrobacter freundii. The PYYDT plasmid backbone is a standard vector used for gene expression in synthetic biology applications. | ||
| − | + | ===Experimental Characterization and results=== | |
Trying to enhance Shewanella’s capacity to produce electricity and polymerize phosphorus, we first introduce PPK1 into it. | Trying to enhance Shewanella’s capacity to produce electricity and polymerize phosphorus, we first introduce PPK1 into it. | ||
Conducting molybdate assays, the level of phosphorus uptake was measured in the Shewanella following the introduction of the plasmid. It was observed that the Shewanella exhibited the most pronounced capacity to polymerize phosphorus following the introduction of PPK1. | Conducting molybdate assays, the level of phosphorus uptake was measured in the Shewanella following the introduction of the plasmid. It was observed that the Shewanella exhibited the most pronounced capacity to polymerize phosphorus following the introduction of PPK1. | ||
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Figure 3: phosphorus accumulation capacity in Shewanella with the introduction of PPK1(SPK1 in the diagram) | Figure 3: phosphorus accumulation capacity in Shewanella with the introduction of PPK1(SPK1 in the diagram) | ||
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Figure 4: electricity production capacity of Shewanella with the introduction of PPK1(SPK1 in the diagram) | Figure 4: electricity production capacity of Shewanella with the introduction of PPK1(SPK1 in the diagram) | ||
SEM and TEM electron microscopy showed that the strains with high phosphorus content had abnormal morphology, indicating that allowing bacteria to be in a high phosphorus content state in advance is not conducive to the rapid transfer of electrons and is not conducive to the ability to produce electricity. | SEM and TEM electron microscopy showed that the strains with high phosphorus content had abnormal morphology, indicating that allowing bacteria to be in a high phosphorus content state in advance is not conducive to the rapid transfer of electrons and is not conducive to the ability to produce electricity. | ||
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Figure 5: Transmission electron microscopy of the Shewanella morphology of WT and modified Shewanella strains with B0034-RBS | Figure 5: Transmission electron microscopy of the Shewanella morphology of WT and modified Shewanella strains with B0034-RBS | ||
| − | + | ===References=== | |
1.Itoh, H., & Shiba, T. (2004). Polyphosphate synthetic activity of polyphosphate:AMP phosphotransferase in Acinetobacter johnsonii 210A. Journal of Bacteriology, 186(15), 5178-5181. | 1.Itoh, H., & Shiba, T. (2004). Polyphosphate synthetic activity of polyphosphate:AMP phosphotransferase in Acinetobacter johnsonii 210A. Journal of Bacteriology, 186(15), 5178-5181. | ||
<!-- Add more about the biology of this part here | <!-- Add more about the biology of this part here | ||
Revision as of 07:02, 29 September 2024
PolyP <->Pi
Contents
Basic Description
This basic part encodes the PPK1 gene which is initially from Citrobacter freundii and we performed codon optimization on, is expressed in the PYYDT plasmid. This basic part is designed to facilitate the reversible conversion between inorganic polyphosphate (PolyP) and inorganic phosphate (Pi). The PPK1 enzyme is known for its ability to synthesize PolyP from ATP and to degrade PolyP back to Pi, with a preference for the synthetic reaction, making it a versatile tool for managing phosphate metabolism in engineered systems. In a sentence, it can reversibly convert Poly p and Pi. This reversible process favors the generation of Poly P.For the first time, we expressed this element in a strain of Shewanella and conducted codon optimization based on Shewanella.
Figure 1: Basic function of PPK1
Construct features(only coding sequence included in basic part)
Promoter: Constitutive promoter for continuous expression. We use tac promoter in our experiment. PPK1 Coding Sequence: Encodes the polyphosphate kinase 1 enzyme. Terminator: Efficient transcription terminator to ensure proper mRNA processing. We use T7Te terminator in our experiment.
Figure 2: Basic construction of PPK1 plasmid
Origin (Organism)
The PPK1 gene was sourced from Citrobacter freundii. The PYYDT plasmid backbone is a standard vector used for gene expression in synthetic biology applications.
Experimental Characterization and results
Trying to enhance Shewanella’s capacity to produce electricity and polymerize phosphorus, we first introduce PPK1 into it. Conducting molybdate assays, the level of phosphorus uptake was measured in the Shewanella following the introduction of the plasmid. It was observed that the Shewanella exhibited the most pronounced capacity to polymerize phosphorus following the introduction of PPK1.
Figure 3: phosphorus accumulation capacity in Shewanella with the introduction of PPK1(SPK1 in the diagram)
Figure 4: electricity production capacity of Shewanella with the introduction of PPK1(SPK1 in the diagram)
SEM and TEM electron microscopy showed that the strains with high phosphorus content had abnormal morphology, indicating that allowing bacteria to be in a high phosphorus content state in advance is not conducive to the rapid transfer of electrons and is not conducive to the ability to produce electricity.
Figure 5: Transmission electron microscopy of the Shewanella morphology of WT and modified Shewanella strains with B0034-RBS
References
1.Itoh, H., & Shiba, T. (2004). Polyphosphate synthetic activity of polyphosphate:AMP phosphotransferase in Acinetobacter johnsonii 210A. Journal of Bacteriology, 186(15), 5178-5181. Sequence and Features
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- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]