Quantitative analysis of JAK binding using isothermal titration calorimetry and surface plasmon resonance

Abstract

Janus Kinases (JAKs) are the key effector kinases that initiate intracellular signalling cascades in response to cytokines and growth factors. As such, a large number of cytoplasmic proteins interact with JAKs both as substrates and as components of regulatory machinery designed to ensure correct activation and termination of signalling. In vitro techniques such as Isothermal Titration Calorimety and Surface Plasmon Resonance are valuable methods to verify and quantify the interaction between JAK and potential binding partners or substrates. Here we describe protocols that exploit both of these in vitro techniques in order to more fully understand the intracellular JAK signalling network.