Iodine deficiency is one of the most common deficiencies in the human body, causing various health problems. Among them, the most common is goiter; but it also causes disturbances in the development of a child and a possibility of spontaneous abortion during pregnancy. The highest amounts of iodine are found in seafood, but in our kitchens it is mostly found in table salt. However, most people do not consume enough of it. The solution for this problem could be biofortification, where sufficiently high amounts of iodine would be introduced into plant products. However, iodine does not represent such an important role for land plants in their organism, which could be a problem that needs to be researched. PorA and CYP18-3 enzymes are found in plants and both play an important role in its development – both are linked to an important role in the photomorphogenesis process. Both proteins are also found in the plant in iodinated form (iodine binds to tyrosines), but it is not known if iodination has any effect on their function.
In order to determine whether the activity of both enzymes changes after iodination, we wanted to prepare it as a recombinant protein. By molecular cloning, we managed to prepare only a transcript for the CYP18-3 protein and insert it into the pET-32a(+) vector. An N-terminal hexahistidine tag and a recognition site for the TEV protease were added to the transcript. We used the bacterial cloning system E. coli DH5α. We were not successful to prepare the protein in the selected bacterial expression system E. coli BL21[DE3] pLysS. It is not clear from our results whether protein expression occurs at all, or whether degradation occurs. The process will still need to be optimized.
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