Src and Syk tyrosine kinases work together in immune signaling and are targets of considerable interest for therapeutics. Immune membrane receptors signal through intracellular sequences, known as ITAMs, that include two tyrosine residues. Src family kinases phosphorylate ITAMs to generate a docking site for Syk through a pair of tandem SH2 domains and initiate a signaling response. This talk will describe molecular dynamics simulations and NMR spectroscopy studies that aim to understand the molecular mechanisms regulating Src and Syk function, particularly recognition of substrate and receptor. MD analysis of the transition pathway for conformational activation of Src kinases finds that helix C rotation presents the greatest energetic barrier to activation, a feature that appears to be commonly exploited in regulation across the kinome. In contrast, Src substrate recognition was found recently from paramagnetic relaxation rates of NMR to differ from other tyrosine kinases while recognition resembles that of threonine/serine kinases. Finally, Syk recognition of ITAM-containing membrane receptors through its tandem SH2 is regulated allosterically through a mechanism that appears to date to be unique to Syk tyrosine kinases; phosphorylation of a tyrosine distal to the ITAM binding sites inhibits receptor binding and releases Syk from the membrane and available for functioning in the cytoplasm and nucleus. This allosteric mechanism has been studied using NMR, ITC and MD and found to involve large conformational changes, but without significant disruption of the binding contacts. Accordingly, the mechanism is essentially all entropy and thus defies the usual entropy/enthalpy compensation that seems to pervade biological associations.