Abstract
Postsynaptic density protein 95 (PSD-95), a specialized scaffold protein with multiple protein interaction domains, forms the backbone of an extensive postsynaptic protein complex that organizes receptors and signal transduction molecules at the synaptic contact zone. Large, detergent-insoluble PSD-95-based postsynaptic complexes can be affinity-purified from conventional PSD fractions using magnetic beads coated with a PSD-95 antibody. In the present study purified PSD-95 complexes were analyzed by LC/MS/MS. A semiquantitative measure of the relative abundances of proteins in the purified PSD-95 complexes and the parent PSD fraction was estimated based on the cumulative ion current intensities of corresponding peptides. The affinity-purified preparation was largely depleted of presynaptic proteins, spectrin, intermediate filaments, and other contaminants prominent in the parent PSD fraction. We identified 525 of the proteins previously reported in parent PSD fractions, but only 288 of these were detected after affinity purification. We discuss 26 proteins that are major components in the PSD-95 complex based upon abundance ranking and affinity co-purification with PSD-95. This subset represents a minimal list of constituent proteins of the PSD-95 complex and includes, in addition to the specialized scaffolds and N-methyl-d-aspartate (NMDA) receptors, an abundance of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors, small G-protein regulators, cell adhesion molecules, and hypothetical proteins. The identification of two Arf regulators, BRAG1 and BRAG2b, as co-purifying components of the complex implies pivotal functions in spine plasticity such as the reorganization of the actin cytoskeleton and insertion and retrieval of proteins to and from the plasma membrane. Another co-purifying protein (Q8BZM2) with two sterile alpha motif domains may represent a novel structural core element of the PSD.
Highlights
Postsynaptic density protein 95 (PSD-95), a specialized scaffold protein with multiple protein interaction domains, forms the backbone of an extensive postsynaptic protein complex that organizes receptors and signal transduction molecules at the synaptic contact zone
postsynaptic density (PSD)-95 complexes were affinity-purified from the conventional PSD fraction using magnetic beads coated with an antibody against the core PSD protein PSD-95
The molecular architecture of the PSD-95 complex is less readily definable than that of the synaptic vesicle as elegantly described by Takamori et al [27]
Summary
Postsynaptic density protein 95 (PSD-95), a specialized scaffold protein with multiple protein interaction domains, forms the backbone of an extensive postsynaptic protein complex that organizes receptors and signal transduction molecules at the synaptic contact zone. We discuss 26 proteins that are major components in the PSD-95 complex based upon abundance ranking and affinity co-purification with PSD-95. The identification of two Arf regulators, BRAG1 and BRAG2b, as copurifying components of the complex implies pivotal functions in spine plasticity such as the reorganization of the actin cytoskeleton and insertion and retrieval of proteins to and from the plasma membrane Another copurifying protein (Q8BZM2) with two sterile ␣ motif domains may represent a novel structural core element of the PSD. In a recent study its total mass was estimated to be around 1 million kDa [1] The function of this massive protein complex appears to be anchoring and organizing postsynaptic neurotransmitter receptors and corresponding signaling molecules at the active zone.
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