Interleukin-4 and IL-10 bind covalently to activated human alpha2-macroglobulin by a mechanism that requires Cys949.
Alpha2-macroglobulin (alpha2M) functions as an extracellular carrier of diverse cytokines, including transforming growth factor-beta1 (TGF-beta1), that expresses anti-inflammatory activities. The results presented here demonstrate that interleukin-10 (IL-10) and IL-4, which also regulate the inflammatory response, bind to alpha2M. Unlike TGF-beta, IL-4 and IL-10 bind almost exclusively to the receptor-recognized, or activated, form of alpha2M. Purified IL-4-alpha2M complexes were predominantly covalent due to thiol disulfide exchange involving Cys949 in the alpha2M subunit. Blocking Cys949 with iodoacetamide significantly inhibited IL-4- and IL-10 binding. Bovine serum albumin (BSA), which possesses a free Cys residue and undergoes thiol disulfide exchange reactions, did not compete with alpha2M for the binding of IL-4 or IL-10. These results suggest a model in which IL-4 and IL-10 associate with activated alpha2M to form complexes that are initially noncovalent but unstable. In these complexes, Cys949 is properly aligned to undergo thiol disulfide exchange and generate stable, covalent IL-4-alpha2M and IL-10-alpha2M complexes.
Garber
T R
TR
Department of Pathology, University of Virginia Health Sciences Center, Charlottesville 22908, USA.
Gonias
S L
SL
Webb
D J
DJ
D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D002417:Cattle; D003432:Cross-Linking Reagents; D003545:Cysteine / Q000737:chemistry; D006801:Humans; D066298:In Vitro Techniques; D016753:Interleukin-10 / Q000737:chemistry / Q000378:metabolism*; D015847:Interleukin-4 / Q000737:chemistry / Q000378:metabolism*; D046911:Macromolecular Substances; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012710:Serum Albumin, Bovine / Q000378:metabolism; D013438:Sulfhydryl Compounds / Q000737:chemistry; D000511:alpha-Macroglobulins / Q000737:chemistry* / Q000378:metabolism*
D003432:Cross-Linking Reagents; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D013438:Sulfhydryl Compounds; D000511:alpha-Macroglobulins; D016753:Interleukin-10; D015847:Interleukin-4; D012710:Serum Albumin, Bovine; D003545:Cysteine