Genome-wide Functional Annotation of Dual-Specificity Protein- and Lipid-Binding Modules that Regulate Protein Interactions

Abstract

Emerging evidence indicates that membrane lipids regulate protein networking by directly interacting with protein-interaction domains (PIDs). As a pilot study to identify and functionally annodate lipid-binding PIDs on a genomic scale, we performed experimental and computational studies of PDZ domains. Characterization of 70 PDZ domains showed that 40% had submicromolar membrane affinity. Using a computational model built from these data, we predicted the membrane binding properties of 2000 PDZ domains from 20 species. The accuracy of the prediction was experimentally validated for 26 PDZ domains. We also subdivided lipid-binding PDZ domains into three classes based on the interplay between membrane and protein binding sites. For different classes of PDZ domains, lipid binding regulates their protein interactions by different mechanisms. Functional studies of a PDZ domain protein, rhophilin2 suggest that all classes of lipid binding PDZ domains serve as genuine dual-specificity modules regulating protein interactions at the membrane under physiological conditions.

Noncanonical Role of the PDZ4 Domain of the Adaptor Protein PDZK1 in the Regulation of the Hepatic High Density Lipoprotein Receptor Scavenger Receptor Class B, Type I (SR-BI)

“…11A (left) show the time courses of binding to the lipid vesicles at the indicated domain concentrations. The sensorgrams show that there was essentially no significant binding of the PDZ3 domain, as reported previously (36), whereas there was a dose-dependent binding of PDZ4. Fig.…”

“…We thus measured the binding of PDZ4 and other PDZ domains from PDZK1 to vesicles whose lipid composition mimics that of the inner leaflet of the plasma membrane of mammalian cells (50). We previously reported that PDZ1, PDZ2, and PDZ3 domains from PDZK1 do not individually bind to such vesicles (36). Here, we measured the binding of individual recombinant PDZ4 and PDZ3 domains to plasma membrane mimetic vesicles.…”

“…Plasma membrane (PM) mimetic vesicles (POPC/1 palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoserine/phosphoinositol/ cholesterol/1,2-dipalmitoyl derivatives of phosphatidylinositol-(4,5)-biphosphonate (12:35:22:8:22:1)) and control 100% POPC vesicles were prepared as described previously (36). All SPR measurements were performed in 20 mM Tris-HCl, pH 7.4, containing 160 mM NaCl at 23°C using a lipid-coated L1 chip in the BIACORE X system as described previously (37).…”

“…Vesicles were injected at 5 l/min onto the corresponding sensor chip surfaces to yield the identical resonance units, ensuring the equal concentration of the coated lipids (PM mimetic and control POPC). Equilibrium measurements were performed at a flow rate of 5 l/min, which allowed enough time for the R values of the association phase to reach near equilibrium levels (R eq ) (36). Each sensorgram was background-corrected by subtracting the control (POPC) surface response from the active (PM mimetic) surface response.…”

“…There are several reports that that PDZ domains can directly bind to membrane lipids (36,48,49) and thus mediate protein-membrane interactions. Such interactions may play an important part in the mechanisms underlying the functions of some PDZ domain-containing proteins.…”