Mechanism of Targeting the a Kinase Anchoring Protein AKAP18δ To the Membrane

Abstract

Many proteins bind phospholipids too weakly to direct membrane association on their own. Localization studies nevertheless, reveal membrane anchoring. Membrane anchoring is furthermore considered to be crucial for their function. AKAP18δ, for example, is part of the signalling cascade which regulates the plasma membrane abundance of the water channel aquaporin-2. The cascade requires both proteins to colocalize in intracellular membranes. In contrast, membrane affinity appears to be rather low as suggested by high sequence homology to the preferentially cytoplasmic AKAP18γ, and the lack of palmitoylation or myristoylation sites which are believed to tailor the homologous proteins AKAP18α and AKAP18β to the membrane. Coincidence detection of a putative binding domain with large net positive charge to negatively charged lipids and specific recognition of a membrane anchored protein (e.g. phosphodiesterase PDE4D) may explain specific membrane targeting of AKAP18δ. Oligomerization of AKAP18δ would also result in an increased membrane affinity by providing several binding sites. To distinguish between both hypotheses, we monitored binding of purified wild type AKAP18δ and AKAP18δ fragments to planar lipid bilayers using fluorescence correlation spectroscopy. Protein binding to both charged and uncharged membranes did not require accessory proteins. Analysis of membrane diffusion constant revealed the existence of oligomers, confirming thereby the second hypothesis.