Differential association of the pleckstrin homology domains of phospholipases C-beta(1), C-beta(2), and C-delta(1) with lipid bilayers and the by subunits of heterotrimeric G proteins

Pleckstrin homology (PH) domains are recognized in more than 100 different proteins, including mammalian phosphoinositide-specific phospholipase C (PL C) isozymes (isotypes beta, gamma, and delta). These structural motifs are thought to function as tethering devices linking their host proteins to mem branes containing phosphoinositides or beta gamma subunits of heterotrimeri c GTP binding (G) proteins. Although the PH domains of PLC-delta and PLC-ga mma have been studied, the comparable domains of the beta isotypes have not . Here, we have measured the affinities of the isolated PH domains of PLC-b eta(1) and -beta(2) (PH-beta(1) and PH-beta(2), respectively) for lipid bil ayers and G-Py subunits. Like the intact enzymes, these PH domains bind to membrane surfaces composed of zwitterionic phosphatidylcholine with moderat e affinity. Inclusion of the anionic lipid phosphatidylserine or phosphatid ylinositol 4,5-bisphosphate [PI(4,5)P-2] and inclusion of G-beta gamma subu nits had little affect on their membrane affinity. In contrast, binding of PLC-delta(1) or its PH domain was highly dependent on PI(4,5)P2. We also de termined whether these domains laterally associate with G-Py subunits bound to membrane surfaces using fluorescence resonance energy transfer. Affinit ies for G-beta gamma were in the following order: PH-beta(2) greater than o r equal to PH-beta(1) > PH-delta(1); the affinities of the native enzyme we re as follows: PLC-beta(2) much greater than PLC-delta(1) > PLC-beta(1). Th us, the PH domain of PLC-beta(1) interacts with G-beta gamma in isolation, but not in the context of the native enzyme. By contrast, docking of the PH domain of PLC-beta(2) with G-beta gamma is comparable to that of the full- length protein and may play a key role in G-beta gamma recognition.