Laboratory for Plant Molecular Biology and Biotechnology, Division of Molecular Biology, Ruđer Bošković Institute, Bijenička cesta 54, 10000 Zagreb, Croatia.
Int J Mol Sci. 2018 Feb 14;19(2):569. doi: 10.3390/ijms19020569.
Thylakoid rhodanase-like protein (TROL) is involved in the final step of photosynthetic electron transport from ferredoxin to ferredoxin: NADP⁺ oxidoreductase (FNR). TROL is located in two distinct chloroplast compartments-in the inner envelope of chloroplasts, in its precursor form; and in the thylakoid membranes, in its fully processed form. Its role in the inner envelope, as well as the determinants for its differential localization, have not been resolved yet. In this work we created six N-terminal amino acid substitutions surrounding the predicted processing site in the presequence of TROL in order to obtain a construct whose import is affected or localization limited to a single intrachloroplastic site. By using in vitro transcription and translation and subsequent protein import methods, we found that a single amino acid exchange in the presequence, Ala67 to Ile67 interferes with processing in the stroma and directs the whole pool of in vitro translated TROL to the inner envelope of chloroplasts. This result opens up the possibility of studying the role of TROL in the chloroplast inner envelope as well as possible consequence/s of its absence from the thylakoids.
类硫氧还蛋白还原酶（TROL）参与从铁氧还蛋白到铁氧还蛋白：NADP⁺氧化还原酶（FNR）的光合作用电子传递的最后一步。TROL 位于两个不同的叶绿体隔室中——叶绿体的内囊体中，以其前体形式存在；和类囊体膜中，以完全加工的形式存在。其在内囊体中的作用以及决定其差异定位的因素尚未得到解决。在这项工作中，我们在 TROL 前导序列的预测加工位点周围创建了六个 N 端氨基酸取代，以获得一种构建体，其导入受到影响或定位仅限于单个类囊体内部。通过使用体外转录和翻译以及随后的蛋白质导入方法，我们发现前导序列中的单个氨基酸交换，Ala67 到 Ile67，干扰了基质中的加工，并将整个体外翻译的 TROL 池引导到叶绿体的内囊体。这一结果为研究 TROL 在叶绿体内膜中的作用以及其从类囊体中缺失的可能后果/影响开辟了可能性。
