Structural analysis of apolipoprotein A-I: Limited proteolysis of methionine-reduced and -oxidized lipid-free and lipid-bound human apo A-I

Academic Article

Abstract

  • The domain structures of lipid-free and lipid-bound apolipoprotein A-I (apo A-I) containing reduced and oxidized methionines were analyzed by limited proteolysis. Lipid-free apo A-I is cleaved primarily in the extreme carboxy-terminus and, to a much lesser extent, in the central region of the protein between residues 115 and 136. Oxidation of methionines 112 and 148 to the corresponding sulfoxides in putative amphipathic helices 4 (P99- E120) and 6 (P143-A164), respectively, causes helices 1 (L44-G65), 2 (P66-S87), and 7 (P165-G186) to become susceptible to protease digestion. These results are consistent with a discrete, globular tertiary structure for the lipid-free protein minimally formed from amphipathic helices 1, 2, 4, 6, and 7. In distinct contrast to lipid-free apo A-I, lipid-bound apo A-I is most susceptible to cleavage in the extreme amino-terminus and, to a lesser extent, in both the central and carboxy-terminal regions. The observed cleavage pattern for the reduced lipid-bound protein supports the existence of many of the turns between helices predicted by sequence analysis of the lipid-bound protein. Methionine oxidation of lipid-bound protein results in a decreased protease susceptibility in the extreme amino-terminus and a concomitant increase in protease susceptibility in the central and carboxy- terminal regions. The results from methionine oxidation indicate the oxidation state of the protein is an important determinant in defining the conformation of both lipid-free and lipid-bound apo A-I.
  • Published In

  • Biochemistry  Journal
  • Digital Object Identifier (doi)

    Author List

  • Roberts LM; Ray MJ; Shih TW; Hayden E; Reader MM; Brouillette CG
  • Start Page

  • 7615
  • End Page

  • 7624
  • Volume

  • 36
  • Issue

  • 24