Sir William Dunn School of Pathology, University of Oxford, Oxford, U.K.
Wolfson Centre for Mathematical Biology, University of Oxford, Oxford, U.K.
Sci Adv. 2017 Mar 24;3(3):e1601692. doi: 10.1126/sciadv.1601692. eCollection 2017 Mar.
Tethered enzymatic reactions are ubiquitous in signaling networks but are poorly understood. A previously unreported mathematical analysis is established for tethered signaling reactions in surface plasmon resonance (SPR). Applying the method to the phosphatase SHP-1 interacting with a phosphorylated tether corresponding to an immune receptor cytoplasmic tail provides five biophysical/biochemical constants from a single SPR experiment: two binding rates, two catalytic rates, and a reach parameter. Tether binding increases the activity of SHP-1 by 900-fold through a binding-induced allosteric activation (20-fold) and a more significant increase in local substrate concentration (45-fold). The reach parameter indicates that this local substrate concentration is exquisitely sensitive to receptor clustering. We further show that truncation of the tether leads not only to a lower reach but also to lower binding and catalysis. This work establishes a new framework for studying tethered signaling processes and highlights the tether as a control parameter in clustered receptor signaling.
连接酶反应在信号网络中无处不在，但了解甚少。本文建立了一种以前未报道的用于表面等离子体共振（SPR）中连接信号反应的数学分析方法。将该方法应用于与磷酸化连接子相互作用的磷酸酶 SHP-1，该连接子对应免疫受体胞质尾部，从单个 SPR 实验中提供了五个生物物理/生化常数：两个结合速率、两个催化速率和一个可达参数。连接绑定通过结合诱导的变构激活（20 倍）和局部底物浓度的显著增加（45 倍）将 SHP-1 的活性提高了 900 倍。可达参数表明，这种局部底物浓度对受体聚集非常敏感。我们还表明，连接子的截断不仅导致可达性降低，而且还导致结合和催化降低。这项工作为研究连接信号过程建立了一个新的框架，并强调了连接子作为聚集受体信号中的控制参数。
