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Sci Adv. 2017 Mar 24;3(3):e1601692. doi: 10.1126/sciadv.1601692. eCollection 2017 Mar.

Biophysical assay for tethered signaling reactions reveals tether-controlled activity for the phosphatase SHP-1.

Author information

1
Sir William Dunn School of Pathology, University of Oxford, Oxford, U.K.
2
Wolfson Centre for Mathematical Biology, University of Oxford, Oxford, U.K.
3
Department of Mathematics and Statistics, Boston University, Boston, MA 02215, USA.
4
Department of Mathematics, University of California, Irvine, Irvine, CA 92697, USA.
5
Sir William Dunn School of Pathology, University of Oxford, Oxford, U.K.; Wolfson Centre for Mathematical Biology, University of Oxford, Oxford, U.K.

Abstract

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.

KEYWORDS:

Biochemistry; SHP-1; allosteric activation; clustered receptors; enzymatic catalysis; mathematical model; stochastic simulations; surface plasmon resonance; tethered signalling; tyrosine phosphatase

PMID:
28378014
PMCID:
PMC5365251
DOI:
10.1126/sciadv.1601692
[Indexed for MEDLINE]
Free PMC Article

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