Department of Life Sciences, Ben-Gurion University of the Negev, Beer Sheva POB 653, Israel.
Int J Mol Sci. 2021 Jan 24;22(3):1136. doi: 10.3390/ijms22031136.
Assembly of photosynthetic complexes is sensitive to changing light intensities, drought and pathogens, each of which induces a redox imbalance that requires the assistance of specific chaperones to maintain protein structure. Here we report a thylakoid membrane-associated DnaJ-like protein, ZnJ6 (Cre06.g251716.t1.2), in . The protein has four CXXCX(G)X(G) motifs that form two zinc fingers (ZFs). Site-directed mutagenesis (Cys > Ser) eliminates the ability to bind zinc. An intact ZF is required for ZnJ6 stability at elevated temperatures. Chaperone assays with recombinant ZnJ6 indicate that it has holding and oxidative activities. ZnJ6 is unable to reduce the disulfide bonds of insulin but prevents its aggregation in a reducing environment. It also assists in the reactivation of reduced denatured RNaseA, possibly by its oxidizing activity. ZnJ6 pull-down assays revealed interactions with oxidoreductases, photosynthetic proteins and proteases. In vivo experiments with a insertional mutant (∆ZnJ6) indicate enhanced tolerance to oxidative stress but increased sensitivity to heat and reducing conditions. Moreover, ∆ZnJ6 has reduced photosynthetic efficiency shown by the Chlorophyll fluorescence transient. Taken together, we identify a role for this thylakoid-associated DnaJ-like oxidizing chaperone that assists in the prevention of protein misfolding and aggregation, thus contributing to stress endurance, redox maintenance and photosynthetic balance.
光合作用复合物的组装对光照强度、干旱和病原体的变化很敏感，每种变化都会导致氧化还原失衡，需要特定伴侣蛋白的协助来维持蛋白质结构。在这里，我们报道了一个类 DnaJ 蛋白，ZnJ6（Cre06.g251716.t1.2），在 中。该蛋白有四个CXXCX(G)X(G)基序，形成两个锌指（ZF）。定点突变（Cys > Ser）消除了结合锌的能力。完整的 ZF 是 ZnJ6 在高温下稳定所必需的。用重组 ZnJ6 进行伴侣蛋白测定表明，它具有结合和氧化活性。ZnJ6 不能还原胰岛素的二硫键，但可以在还原环境中防止其聚集。它还可以协助还原变性 RNaseA 的复性，可能是通过其氧化活性。ZnJ6 的下拉实验显示与氧化还原酶、光合作用蛋白和蛋白酶的相互作用。带有 插入突变体（∆ZnJ6）的体内实验表明，该突变体对氧化应激的耐受性增强，但对热和还原条件的敏感性增加。此外，∆ZnJ6 的光合作用效率降低，表现在叶绿素荧光瞬变上。总之，我们确定了一个类 DnaJ 相关的叶绿体伴侣蛋白的氧化还原功能，它可以协助防止蛋白质错误折叠和聚集，从而有助于耐受应激、维持氧化还原平衡和光合作用。
