Rial Daniela V, Lombardo Verónica A, Ceccarelli Eduardo A, Ottado Jorgelina
Molecular Biology Division, IBR (Instituto de Biología Molecular y Celular de, Rosario), CONICET, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Rosario, Argentina.
Eur J Biochem. 2002 Nov;269(22):5431-9. doi: 10.1046/j.1432-1033.2002.03233.x.
Protein transport across organelles' membranes requires that precursor proteins adopt an unfolded structure in order to be translocated by the import machinery. Ferredoxin-NADP+ reductase precursor, as well as many others, acquires a tightly folded structure that needs to be unfolded before or during its import. Several steps of chloroplast protein import are not fully understood. In particular, the role of different regions of the precursor protein has not been completely elucidated. In this work, we have studied the import into chloroplasts of precursor proteins with inclusions of amino acid spacers between the transit peptide and the mature protein, and with deletions in the N-terminal region of the mature enzyme. We measured the import rate constants for these precursors and the results indicate that the distance between the transit peptide and the core of the mature protein determines the import kinetics. The longer precursors were imported into the organelle faster than the wild type form. Precursors with deletions in the N-terminal region of the mature protein also showed increased import rates compared to the wild type. Homology studies amongst all family members reveal that only chloroplastic proteins possess this region. We suggest that even if the first amino acids of the mature protein do not contribute to its overall structural stability, they condition the kinetic parameters of the import reaction. Besides, the distance between the transit peptide and the mature protein core may be modulating the import rate at which the chloroplast incorporates this protein from the cytosol.
蛋白质跨细胞器膜的转运要求前体蛋白采用未折叠的结构，以便被导入机制转运。铁氧还蛋白-NADP⁺还原酶前体以及许多其他前体蛋白都具有紧密折叠的结构，在导入之前或导入过程中需要展开。叶绿体蛋白质导入的几个步骤尚未完全了解。特别是，前体蛋白不同区域的作用尚未完全阐明。在这项工作中，我们研究了在转运肽和成熟蛋白之间包含氨基酸间隔区以及成熟酶N端区域有缺失的前体蛋白导入叶绿体的情况。我们测量了这些前体蛋白的导入速率常数，结果表明转运肽与成熟蛋白核心之间的距离决定了导入动力学。较长的前体蛋白比野生型形式更快地导入细胞器。成熟蛋白N端区域有缺失的前体蛋白与野生型相比，导入速率也有所增加。对所有家族成员的同源性研究表明，只有叶绿体蛋白具有该区域。我们认为，即使成熟蛋白的前几个氨基酸对其整体结构稳定性没有贡献，但它们会影响导入反应的动力学参数。此外，转运肽与成熟蛋白核心之间的距离可能在调节叶绿体从细胞质中摄取该蛋白的导入速率。
