Heterologous Reconstitution of the Biosynthesis Pathway for Producing Holo-C-Phycocyanin Alpha Subunit (CpcA) from...

C-phycocyanin (C-PC), a blue protein pigment found in cyanobacteria, has potential properties for use in food and medical applications, including antioxidants, antimicrobial, and anti-inflammatory. Generally, C-PC is extracted from mesophilic cyanobacteria, such as Arthrospira platensis, which is unstable under high-temperature conditions. Therefore, thermophilic cyanobacteria represent potential sources of thermostable C-PC. This study aimed to produce a holo-C-PC alpha subunit (holo-CpcA) from the thermophilic cyanobacterium Thermosynechococcus sp. TUBT-T01 by reconstituting the pathway in Escherichia coli. The constructed expression plasmid was designed with a polycistronic transcription unit containing ho1, pcyA, cpcE, and cpcF inserted into pET3a. Together with the pET28a carrying cpcA gene, two plasmids were co-transformed and co-expressed in E. coli Rosetta (DE3). Following induction, the cell pellet exhibited a visible color change, suggesting successful expression of the C-PC alpha subunit. The antioxidant activity of crude holo-CpcA was analyzed; it showed higher antioxidant activity compared to crude apo-CpcA in both the ABTS radical cation decolorization assay and the ORAC-Fluorescein Assay. This study reported the successful heterologous production of holo-CpcA from Thermosynechococcus sp. in E. coli, and the protein also demonstrated antioxidant activities.