The broad range nucleotide transporter PtNTT2 from Phaeodactylum tricornutum facilitates the counter exchange of nucleotide triphosphates (NTPs) and nucleoside diphosphates (NDPs), importing NTPs into the plastid. Expressed in E. coli, it allows the uptake of various nucleotides from the cultivation media. This function is essential for the development of a semisynthetic organism, which uses externally provided unnatural nucleotides for replication of its semisynthetic DNA. For our project, we characterized the structure, kinetics, dynamics and subcellular localization of PtNTT2 and different PtNTT2 variants. Therefore, we applied bioinformatic prediction tools, confocal laser scanning microscopy, SDS-PAGE, Western Blot, MALDI-TOF as well as HPLC-MS. All PtNTT2 variants were also cultivated to investigate the effect of PtNTT2 expression on the growth rate. In the future, advanced endosymbiotic systems or novel biosafety mechanisms can be developed based on this part.
