title: [Binding Characterizations of Asp-hemolysin to Oxidized Low-Density Lipoprotein].
abstract: Oxidized low-density lipoprotein (Ox-LDL) is known to be involved in the generation and progression of atherosclerosis. Ox-LDL has a number of potentially atherogenic effects on vascular cells, including uncontrol uptake by scavenger receptors. Asp-hemolysin, a hemolytic toxin from Aspergillus fumigatus, is a binding protein for Ox-LDL. This study was undertaken to clarify the binding specificity of Asp-hemolysin to Ox-LDL. We examined the binding specificity of Asp-hemolysin to Ox-LDL using several modified lipoproteins and scavenger-receptor ligands. Asp-hemolysin bound to Ox-LDL with shorter LDL oxidation times. However, Asp-hemolysin did not bind to acetylated LDL. The native high-density lipoprotein (n-HDL) and modified HDL (e.g., acetylated HDL, oxidized HDL) also had no Asp-hemolysin binding. Inhibitors of scavenger-receptor binding, including maleylated bovine serum albumin, polyinosinic acid, dextran sulfate, and fucoidin, had no effect on the binding of Ox-LDL to Asp-hemolysin. Surface plasmon-resonance studies revealed that Ox-LDL binds with high affinity (K(D)=0.63 microg/ml) to Asp-hemolysin. Furthermore, we have shown that Ox-LDL strongly inhibits the hemolytic activity of Asp-hemolysin and that the removal of lysophosphatidylcholine (lysoPC) from Ox-LDL abolished the inhibition. We also investigated the interaction between Asp-hemolysin and lysoPC as a typical lipid moiety of Ox-LDL. The binding of Asp-hemolysin to LDL oxidized for different times depended on the lysoPC content in each Ox-LDL. In addition, the inhibition of lysoPC production in Ox-LDL by phenylmethylsulfonyl fluoride (PMSF) pretreatment of LDL resulted in a marked decrease in Asp-hemolysin binding to PMSF-pretreated Ox-LDL. The binding analysis of Asp-hemolysin to lysoPC revealed that Asp-hemolysin binds directly to lysoPC. We conclude that Asp-hemolysin is a specific binding protein with high affinity for Ox-LDL and that its binding specificity is distinct from any receptor for Ox-LDL.
authors.affiliation: Sendai Hospital of East Japan Railway Company, Sendai, Japan. senyaku@jr-hospital.aoba.sendai.jp
authors.forename: Yoichi
authors.initials: Y
authors.lastname: Kudo
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001161:Arteriosclerosis / Q000209:etiology; D001232:Aspergillus fumigatus; D001483:Base Sequence; D001665:Binding Sites; D006460:Hemolysin Proteins / Q000378:metabolism*; D006801:Humans; D008077:Lipoproteins, LDL / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008244:Lysophosphatidylcholines / Q000378:metabolism; D009183:Mycotoxins / Q000378:metabolism*; D011485:Protein Binding
chemical_list: D006460:Hemolysin Proteins; D008077:Lipoproteins, LDL; D008244:Lysophosphatidylcholines; D009183:Mycotoxins; C099252:oxidized low density lipoprotein