title: Structural Stability and Local Dynamics in Disease-Causing Mutants of Human Apolipoprotein A-I: What Makes the Protein Amyloidogenic?
abstract: ApoA-I, the major protein of plasma high-density lipoprotein, removes cellular cholesterol and protects against atherosclerosis. ApoA-I mutations can cause familial amyloidosis, a life-threatening disease wherein N-terminal protein fragments form fibrils in vital organs. To unveil the protein misfolding mechanism and to understand why some mutations cause amyloidosis while others do not, we analyzed the structure, stability, and lipid-binding properties of naturally occurring mutants of full-length human apoA-I causing either amyloidosis (G26R, W50R, F71Y, and L170P) or aberrant lipid metabolism (L159R). Global and local protein conformation and dynamics in solution were assessed by circular dichroism, fluorescence, and hydrogen-deuterium exchange mass spectrometry. All mutants showed increased deuteration in residues 14-22, supporting our hypothesis that decreased protection of this major amyloid "hot spot" can trigger protein misfolding. In addition, L159R showed local helical unfolding near the mutation site, consistent with cleavage of this mutant in plasma to generate the labile 1-159 fragment. Together, the results suggest that reduced protection of the major amyloid "hot spot", combined with the structural integrity of the native helix bundle conformation, shifts the balance from protein clearance to β-aggregation. A delicate balance between the overall structural integrity of a globular protein and the local destabilization of its amyloidogenic segments may be a fundamental determinant of this and other amyloid diseases. Furthermore, mutation-induced conformational changes observed in the helix bundle, which comprises the N-terminal 75% of apoA-I, and its flexible C-terminal tail suggest the propagation of structural perturbations to distant sites via an unexpected template-induced ensemble-based mechanism, challenging the classical structure-based view.
authors.affiliation: Department of Physiology & Biophysics, Boston University School of Medicine, 700 Albany Street, Boston, MA 02118, USA.
authors.forename: Madhurima
authors.initials: M
authors.lastname: Das
authors.affiliation: Department of Chemistry & Chemical Biology, Northeastern University, 360 Huntington Avenue, Boston, MA 02115, USA.
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Wilson
authors.affiliation: Department of Physiology & Biophysics, Boston University School of Medicine, 700 Albany Street, Boston, MA 02118, USA.
authors.forename: Xiaohu
authors.initials: X
authors.lastname: Mei
authors.affiliation: Department of Chemistry & Chemical Biology, Northeastern University, 360 Huntington Avenue, Boston, MA 02115, USA.
authors.forename: Thomas E
authors.initials: TE
authors.lastname: Wales
authors.affiliation: Department of Chemistry & Chemical Biology, Northeastern University, 360 Huntington Avenue, Boston, MA 02115, USA.
authors.forename: John R
authors.initials: JR
authors.lastname: Engen
authors.affiliation: Department of Physiology & Biophysics, Boston University School of Medicine, 700 Albany Street, Boston, MA 02118, USA. Electronic address: Gursky@bu.edu.
authors.forename: Olga
authors.initials: O
authors.lastname: Gursky
mesh_terms: D000682:Amyloid / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016632:Apolipoprotein A-I / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002942:Circular Dichroism; D006801:Humans; D050356:Lipid Metabolism; D013058:Mass Spectrometry; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D056004:Molecular Dynamics Simulation; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D066263:Protein Aggregation, Pathological; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D055550:Protein Stability
chemical_list: D000682:Amyloid; D016632:Apolipoprotein A-I; D050505:Mutant Proteins
keywords: familial amyloidosis and atherosclerosis; hydrogen–deuterium exchange mass spectrometry; protein stability and misfolding; protein–lipid interactions; transmission of conformational changes