Membrane binding and clustering of NAP-22, a neuron-enriched intrinsically disordered protein

Neuronal acidic protein of 22 kDa (NAP-22, also called CAP23 or BASP1) is a myristoylated protein highly expressed in brain (> 0.4% of the total protein), and it highly localizes in the presynaptic region. NAP-22 was found to be a major component of detergent-resistant membrane fraction of brain, indicating its affinity to the cholesterol-enriched lipid domains (lipid rafts). In addition to several proteins, NAP-22 was found to interact with various membrane lipids, and these results suggested a variety roles of this protein in the membrane dynamics and signal transduction in neurons. Here, we studied the membrane binding of NAP-22 using a substrate-supported lipid bilayer having well-defined lipid compositions and geometries. We found that myristoylation of NAP-22 was critical for its membrane binding. Acidic lipids such as phosphatidylserine and phosphatidylinositol 4,5-bisphosphate promoted the membrane binding, but cholesterol had no effect. Interaction of NAP-22 and lipid raft was assessed using a model membrane with patterned liquid-ordered and liquid-disordered regions. NAP-22 predominantly partitioned in liquid-disordered region, but affinity to liquid-ordered region was enhanced by acidic lipids. On the other hand, binding of NAP-22 to brain-derived detergent-resistant membrane fraction was diminished by the protease treatment. These results indicate that electrostatic and protein-protein interactions play major roles in the accumulation of NAP-22 in DRM. Furthermore, clustering of NAP-22 was observed only on the membrane containing acidic lipid (phosphatidylserine), suggesting an intricate relation between membrane binding and clustering, which should play profound roles in the physiological functions of NAP-22 in neurons.