PT - JOURNAL ARTICLE AU - Jesse Goyette AU - Citlali Solis Salas AU - Nicola Coker-Gordon AU - Marcus Bridge AU - Samuel A. Isaacson AU - Jun Allard AU - Omer Dushek TI - Biophysical assay for tethered signaling reactions reveals tether-controlled activity for the phosphatase SHP-1 AID - 10.1101/063776 DP - 2016 Jan 01 TA - bioRxiv PG - 063776 4099 - http://biorxiv.org/content/early/2016/07/14/063776.short 4100 - http://biorxiv.org/content/early/2016/07/14/063776.full AB - Tethered enzymatic reactions are ubiquitous in signalling networks but are poorly understood. Here, a novel mathematical analysis is established for tethered signalling 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 5 biophysical/biochemical constants from a single SPR experiment: two binding rates, two catalytic rates, and a reach parameter. Tether binding increased the activity of SHP-1 by 900-fold through a binding-induced allosteric activation (20-fold) and a more significant increase in local sub-strate 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. The work establishes a new framework for studying tethered signalling processes and highlights the tether as a control parameter in clustered signalling.