title: Murine apolipoprotein serum amyloid A in solution forms a hexamer containing a central channel.
abstract: Serum amyloid A (SAA) is a small apolipoprotein that binds to high-density lipoproteins in the serum. Although SAA seems to play a role in host defense and lipid transport and metabolism, its specific functions have not been defined. Despite the growing implications that SAA plays a role in the pathology of various diseases, a high-resolution structure of SAA is lacking because of limited solubility in the high-density lipoprotein-free form. In this study, complementary methods including glutaraldehyde cross-linking, size-exclusion chromatography, and sedimentation-velocity analytical ultracentrifugation were used to show that murine SAA2.2 in aqueous solution exists in a monomer-hexamer equilibrium. Electron microscopy of hexameric SAA2.2 revealed that the subunits are arranged in a ring forming a putative central channel. Limited trypsin proteolysis and mass spectrometry analysis identified a significantly protease-resistant SAA2.2 region comprising residues 39-86. The isolated 39-86 SAA2.2 fragment did not hexamerize, suggesting that part of the N terminus is involved in SAA2.2 hexamer formation. Circular-dichroism spectrum deconvolution and secondary-structure prediction suggest that SAA2.2 contains approximately 50% of its residues in alpha-helical conformation and <10% in beta-structure. These findings are consistent with the recent discovery that human SAA1.1 forms a membrane channel and have important implications for understanding the 3D structure, multiple functions, and pathological roles of this highly conserved protein.
authors.affiliation: Department of Chemistry, Rensselaer Polytechnic Institute, 110 8th Street, Troy, NY 12180, USA.
authors.forename: Limin
authors.initials: L
authors.lastname: Wang
authors.forename: Hilal A
authors.initials: HA
authors.lastname: Lashuel
authors.forename: Thomas
authors.initials: T
authors.lastname: Walz
authors.forename: Wilfredo
authors.initials: W
authors.lastname: Colon
mesh_terms: D000686:Amyloidosis / Q000378:metabolism; D000818:Animals; D001704:Biopolymers; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D003432:Cross-Linking Reagents / Q000494:pharmacology; D005976:Glutaral / Q000494:pharmacology; D013058:Mass Spectrometry; D051379:Mice; D008854:Microscopy, Electron; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D000685:Serum Amyloid A Protein / Q000737:chemistry*; D013329:Structure-Activity Relationship; D014357:Trypsin / Q000494:pharmacology; D014461:Ultracentrifugation
chemical_list: D001704:Biopolymers; D003432:Cross-Linking Reagents; D011993:Recombinant Fusion Proteins; C489995:Saa2 protein, mouse; D000685:Serum Amyloid A Protein; D014357:Trypsin; D005976:Glutaral