title: Denaturation and self-association of apolipoprotein A-I investigated by electrophoretic techniques.
abstract: Purified human apolipoprotein A-I (apoA-I), when run across a transverse urea gradient at alkaline pH, gives a complex pattern characterized by a number of parallel sigmoidal curves, in which the transition between high- and low-mobility forms, i.e. from folded to unfolded structure, occurs between 1.1 and 3.2 M urea. Size differences appear to be the major cause of this isomerism. When migrated across a wide pH range in the presence of varying amounts of urea to display its titration curve, apoA-I is resolved into two pairs of bands, running parallel in the neutral to basic pH region while merging at acidic pH; such a finding does not correlate with a differential exposure of His residues, as shown by diethyl pyrocarbonate titration. Ferguson plot analysis, confirmed by cross-linking experiments, demonstrates a gradual shift from higher to lower mass aggregates as the urea concentration is raised; the monomeric form undergoes denaturation by swelling to an approximately 50% larger hydrodynamic volume than in its native state. At alkaline pH, where apoA-I exists as aggregated species, disaggregation and unfolding appear to happen at once, the larger aggregates being less stable than the smaller ones. At acidic pH, apoA-I does not form aggregates and has little secondary structure; unfolding is then a progressive rather than a cooperative process.
authors.affiliation: Istituto di Scienze Farmacologiche and Centro Enrica Grossi Paoletti, Facoltà di Farmacia, Università di Milano, Italy. gianazza@isfunix.farma.unimi.it
authors.forename: E
authors.initials: E
authors.lastname: Gianazza
authors.forename: L
authors.initials: L
authors.lastname: Calabresi
authors.forename: O
authors.initials: O
authors.lastname: Santi
authors.forename: C R
authors.initials: CR
authors.lastname: Sirtori
authors.forename: G
authors.initials: G
authors.lastname: Franceschini
mesh_terms: D016632:Apolipoprotein A-I / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D011489:Protein Denaturation
chemical_list: D016632:Apolipoprotein A-I