Proteins encoded by the sll0217-sll0218-sll0219 operon in cyanobacterium Synechocystis sp. PCC 6803
Ruohisto, Essi (2018-02-13)
Proteins encoded by the sll0217-sll0218-sll0219 operon in cyanobacterium Synechocystis sp. PCC 6803
Ruohisto, Essi
(13.02.2018)
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Turun yliopisto
Tiivistelmä
It has been shown that the proteins encoded by the flv4-2 operon are crucial in the photoprotection of Photosystem II. However, the photoprotective mechanism operated by these proteins still remains unknown. The aim of our study was to elucidate the interaction partners of the flv4-2 operon-encoded proteins by purifying the proteins with affinity chromatography. For that purpose the C- and/or N- termini of the proteins were fused with an affinity tag. The tagged flv2-4 operon constructs were introduced to the chromosome of a cyanobacterium, Synechocystis sp. PCC 6803.
The fusion proteins were purified with affinity resins which specifically bind the tags fused to the proteins. In our hands the purification of the tagged Flv2 and Flv4 proteins proved to be inefficient with the standard affinity chromatographic methods. We speculate that the attached tags were enclosed inside the Flv2/Flv4 heterodimer, and thus binding to the affinity resin was obstructed. Nevertheless, we were able to detect some protein bands that were specific to the elution fraction obtained from purified FLv4 in comparison to wild type control. These proteins might be interacting partners of the Flv2 and Flv4, however, they need to be further studied. Purification of Sll0218 was more successful, but interaction partners of this protein remain to be elucidated.
We conclude that some of the used affinity chromatography methods proved to be promising for purification of tagged flv4-2 operon-encoded proteins. However, reliable results and efficient purification of the fusion proteins require careful optimisation of the chosen methods or modifications to the tagged flv2-4 constructs.
The fusion proteins were purified with affinity resins which specifically bind the tags fused to the proteins. In our hands the purification of the tagged Flv2 and Flv4 proteins proved to be inefficient with the standard affinity chromatographic methods. We speculate that the attached tags were enclosed inside the Flv2/Flv4 heterodimer, and thus binding to the affinity resin was obstructed. Nevertheless, we were able to detect some protein bands that were specific to the elution fraction obtained from purified FLv4 in comparison to wild type control. These proteins might be interacting partners of the Flv2 and Flv4, however, they need to be further studied. Purification of Sll0218 was more successful, but interaction partners of this protein remain to be elucidated.
We conclude that some of the used affinity chromatography methods proved to be promising for purification of tagged flv4-2 operon-encoded proteins. However, reliable results and efficient purification of the fusion proteins require careful optimisation of the chosen methods or modifications to the tagged flv2-4 constructs.