Manak Michael S, Ferl Robert J
Program in Plant Molecular and Cellular Biology, Horticultural Sciences, University of Florida, 1143 Hull Road, Fifield Hall 110690, Gainesville, Florida 32611-0690, USA.
Biochemistry. 2007 Jan 30;46(4):1055-63. doi: 10.1021/bi061366c.
Oscillations in cellular divalent cation concentrations are key events that can trigger signal transduction cascades. Common cellular divalent cations, such as calcium and magnesium, interact with 14-3-3 proteins. The metal ion interaction causes a conformational change in the 14-3-3 proteins, which is manifested as an increase in hydrophobicity. In this study, the effect of divalent cations on the interaction between 14-3-3 proteins and target peptides was investigated using surface plasmon resonance and isothermal titration calorimetry. The binding between ten recombinant Arabidopsis 14-3-3 isoforms and two synthetic target peptides was observed in the presence of various physiologically relevant concentrations of calcium or magnesium, from 1 microM to 1 mM or from 1 microM to 5 mM, respectively. The synthetic target peptides were based on sequences from Arabidopsis nitrate reductase (NR2) and the plasma membrane proton pump (AHA2) representing fundamentally different target classes. Isoforms representing every branch of the Arabidopsis 14-3-3 phylogenetic tree were tested. The general result for all cases is that an increased concentration of divalent cations in solution causes an increase in the concentration of 14-3-3 protein interacting with the respective phosphopeptide.
细胞中二价阳离子浓度的振荡是能够触发信号转导级联反应的关键事件。常见的细胞二价阳离子，如钙和镁，与14-3-3蛋白相互作用。金属离子相互作用会导致14-3-3蛋白发生构象变化，表现为疏水性增加。在本研究中，使用表面等离子体共振和等温滴定量热法研究了二价阳离子对14-3-3蛋白与靶肽之间相互作用的影响。在分别存在1微摩尔至1毫摩尔或1微摩尔至5毫摩尔的各种生理相关浓度的钙或镁的情况下，观察到了十种重组拟南芥14-3-3亚型与两种合成靶肽之间的结合。合成靶肽基于拟南芥硝酸还原酶（NR2）和质膜质子泵（AHA2）的序列，代表了根本不同的靶标类别。测试了代表拟南芥14-3-3系统发育树每个分支的亚型。所有情况下的总体结果是，溶液中二价阳离子浓度的增加会导致与相应磷酸肽相互作用的14-3-3蛋白浓度增加。
