Part:BBa_K5034222
Pi <-> Poly P
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.
Construct features
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.
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
Alteration of protein expression intensity can regulate the metabolic networks, so we focused on RBS with varying translation strengths to facilitate the regulation of PPK1 concentration in Shewanella thus developing the best ability to produce electricity and polymerize phosphorus. Using molybdate assays to determine Pi concentration and half-cell reaction(electrochemistry) to measure the electricity production ability, we found SPK3(with RBS BBa-B0034) has the greatest capacity to polymerize phosphorus but a worst electroproduction capability.
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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