The binding of FNR to PSI has been postulated long ago, however, a clear evidence is still missing. In this work, using isothermal titration calorimetry (ITC), we found that FNR binds to photosystem I with its light harvesting complex I (PSI-LHCI) from C. reinhardtii with a 1:1 stoichiometry, a Kd of ~0.8 μM and ∆H of -20.7 kcal/mol. Titrations at different temperatures were used to determine the heat capacity change, ∆CP, of the binding, through which the size of the interface area between the proteins was assessed as ~3000 Å2. In a different set of ITC experiments, introduction of various sucrose concentrations was used to estimate that ~95 water molecules are released to the solvent. These observations support the notion of a binding site shared by few of the photosystem I - light harvesting complex I (PSI-LHCI) subunits in addition to PsaE. Based on these results, a hypothetical model was built for the binding site of FNR at PSI, using known crystallographic structures of: cyanobacterial PSI in complex with ferredoxin (Fd), plant PSI-LHCI and Fd:FNR complex from cyanobacteria. FNR binding site location is proposed to be at the foot of the stromal ridge and above the inner LHCI belt. It is expected to form contacts with PsaE, PsaB, PsaF and at least one of the LHCI. In addition, a ~4.5-fold increased affinity between FNR and PSI-LHCI under crowded 1 M sucrose environment led us to conclude that in C. reinhardtii FNR also functions as a subunit of PSI-LHCI.

中文翻译：

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铁氧还蛋白NADP +氧化还原酶（FNR）与植物光系统I的结合

FNR与PSI的绑定早就被假定，但是，仍然缺少明确的证据。在这项工作中，我们使用等温滴定热法（ITC），发现FNR以1：1的化学计量比，约0.8μM的Kd和∆与来自雷氏梭菌的光收集复合体I（PSI-LHCI）结合到光系统I。 H为-20.7kcal / mol。使用不同温度下的滴定来确定结合物的热容变化ΔCP，据此，蛋白质之间的界面区域大小约为3000Å​​2。在一组不同的ITC实验中，采用了各种浓度的蔗糖来估计约95个水分子被释放到溶剂中。这些观察结果支持了结合位点的概念，该结合位点由PsaE以外的几个光系统I-光捕获复合体I（PSI-LHCI）亚基共享。基于这些结果，使用以下已知的晶体结构，建立了FNR在PSI上的结合位点的假想模型：蓝细菌PSI与铁氧还蛋白（Fd）的复合体，植物PSI-LHCI和蓝藻的Fd：FNR复合体。FNR结合位点的位置建议在基质脊的底部和内部LHCI带的上方。预计将与PsaE，PsaB，PsaF和LHCI中的至少一种形成接触。此外，在拥挤的1 M蔗糖环境下，FNR和PSI-LHCI之间的亲和力增加了约4.5倍，这使我们得出结论，在莱茵衣藻中，FNR也是PSI-LHCI的一个亚基。来自蓝细菌的FNR复合物。FNR结合位点的位置建议在基质脊的底部和内部LHCI带的上方。预计将与PsaE，PsaB，PsaF和LHCI中的至少一种形成接触。此外，在拥挤的1 M蔗糖环境下，FNR和PSI-LHCI之间的亲和力增加了约4.5倍，这使我们得出结论，在莱茵衣藻中，FNR也是PSI-LHCI的一个亚基。来自蓝细菌的FNR复合物。FNR结合位点的位置建议在基质脊的底部和内部LHCI带的上方。预计将与PsaE，PsaB，PsaF和LHCI中的至少一种形成接触。此外，在拥挤的1 M蔗糖环境下，FNR和PSI-LHCI之间的亲和力增加了约4.5倍，这使我们得出结论，在莱茵衣藻中，FNR也是PSI-LHCI的一个亚基。