Plant phytochromes enable vital adaptations to red and far-red light. At the molecular level, these responses are mediated by light-regulated interactions between phytochromes and partner proteins, foremost the phytochrome-interacting factors (PIF). Although known for decades, quantitative analyses of these interactions have long been sparse. To address this deficit, we here studied by an integrated fluorescence-spectroscopic approach the equilibrium and kinetics of Arabidopsis thaliana phytochrome B binding to a tetramerized PIF6 variant. Several readouts consistently showed the stringently light-regulated interaction to be little affected by PIF tetramerization. Analysis of the binding kinetics allowed the determination of bimolecular association and unimolecular dissociation rate constants as a function of light. Unexpectedly, the stronger affinity of A. thaliana phytochrome B under red light relative to far-red light is entirely due to accelerated association rather than decelerated dissociation. The association reaction under red light is highly efficient and only 3-fold slower than the diffusion limit. The present findings pertain equally to the analysis of signal transduction in plants and to the biotechnological application of phytochromes.

植物光敏色素能够对红光和远红光进行重要的适应。在分子水平上，这些反应是由光敏色素和伴侣蛋白之间的光调节相互作用介导的，最重要的是光敏色素相互作用因子 (PIF)。尽管已知数十年，但长期以来很少对这些相互作用进行定量分析。为了解决这一缺陷，我们在这里通过综合荧光光谱法研究了拟南芥光敏色素 B的平衡和动力学与四聚化 PIF6 变体结合。几个读数始终显示严格光调节的相互作用受 PIF 四聚化的影响很小。结合动力学分析允许确定双分子结合和单分子解离速率常数作为光的函数。出乎意料的是，A. thaliana 光敏色素 B在红光下相对于远红光具有更强的亲和力，这完全是由于加速结合而不是减速解离。红光下的缔合反应效率很高，仅比扩散极限慢 3 倍。目前的发现同样适用于植物信号转导分析和光敏色素的生物技术应用。