A heat shock protein70 fusion protein with alpha1-antitrypsin in plasma of type 1 diabetic subjects.
The recent observation that heat shock proteins (HSPs), mostly glucose regulated protein94 (Grp94) and HSP70, are present in plasma of Type 1 diabetic subjects as complexes with immunoglobulins, prompted us to investigate the nature and extent of this association, whether it represents HSP-induced activation of the immune system. Two complementary affinity chromatography procedures followed by immunoprecipitation and immunoblot analyses of HSP-enriched, plasma-purified peaks, revealed that HSPs were inextricably linked with IgG in SDS-resistant complexes from which proteins dissociate partially under reducing treatment. HSP70 was found also closely linked with alpha1-antitrypsin (alpha1AT) in a single protein having the mass of alpha1AT but elution characteristics different from those of normal alpha1AT. Immunoprecipitation with anti-HSP70 antibodies led to co-immunoprecipitation of the alpha1AT species linked to HSP70, thus confirming fusion of the proteins. The additional finding of circulating antibodies against the HSP70-alpha1AT protein supported its immunogenic properties with implications for diabetes and its complications.
Finotti
Paola
P
Department of Pharmacology and Anaesthesiology, University of Padua, Largo E. Meneghetti 2, 35131 Padua, Italy. paola.finotti@unipd.it
Pagetta
Andrea
A
D000328:Adult; D015153:Blotting, Western; D002846:Chromatography, Affinity; D003922:Diabetes Mellitus, Type 1 / Q000097:blood*; D004305:Dose-Response Relationship, Drug; D004586:Electrophoresis; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D018840:HSP70 Heat-Shock Proteins / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D007328:Insulin / Q000378:metabolism; D008297:Male; D008565:Membrane Proteins / Q000737:chemistry; D008875:Middle Aged; D010949:Plasma / Q000378:metabolism*; D011233:Precipitin Tests; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000737:chemistry* / Q000378:metabolism; D012685:Sepharose / Q000737:chemistry; D000515:alpha 1-Antitrypsin / Q000737:chemistry* / Q000378:metabolism
D018840:HSP70 Heat-Shock Proteins; D007328:Insulin; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; D000515:alpha 1-Antitrypsin; C018551:glucose-regulated proteins; D012685:Sepharose