Pilon M, Rietveld A G, Weisbeek P J, de Kruijff B
Institute of Molecular Biology and Medical Biotechnology, University of Utrecht, The Netherlands.
J Biol Chem. 1992 Oct 5;267(28):19907-13.
Ferredoxin is a chloroplast stroma protein which is cytosolically synthesized as a precursor with an amino-terminal extension called the transit sequence that is needed for the post-translational uptake by the chloroplast. To characterize the secondary and tertiary structure elements, the full precursor, the holo- and apo- (without iron-sulfur cluster) forms of the mature protein, and the chemically synthesized transit peptide were obtained and analyzed separately. Circular dichroism, tryptophan fluorescence quenching, and protease accessibility experiments indicate that the precursor has a low content of defined secondary structure and resembles unfolded proteins; these properties are due to both the mature part and the transit sequence. This result provides an explanation for the lack of cytosolic factor requirement of this protein for import. In an import competition assay, the isolated transit peptide had an affinity for the chloroplasts comparable to the full precursor. Interestingly and of possible importance to the import process, the transit peptide has conformational flexibility as it adopts alternative secondary structures in different environments.
铁氧化还原蛋白是一种叶绿体基质蛋白，它在细胞质中以前体形式合成，前体带有一个氨基末端延伸序列，称为转运肽序列，这是叶绿体进行翻译后摄取所必需的。为了表征二级和三级结构元件，分别获得并分析了完整前体、成熟蛋白的全铁形式和脱辅基（无铁硫簇）形式以及化学合成的转运肽。圆二色性、色氨酸荧光猝灭和蛋白酶可及性实验表明，前体具有低含量的确定二级结构，类似于未折叠的蛋白质；这些特性是由成熟部分和转运肽序列共同导致的。这一结果解释了该蛋白导入过程中对细胞质因子需求的缺乏。在导入竞争试验中，分离出的转运肽对叶绿体的亲和力与完整前体相当。有趣的是，对于导入过程可能很重要的是，转运肽具有构象灵活性，因为它在不同环境中会采用不同的二级结构。
