A preliminary study for isolation of catalytic antibodies by histidine ligand affinity chromatography as an alternative to conventional protein A/G methods.
Catalytic autoimmune antibodies from the sera of lupus patients were purified using histidyl-aminohexyl-Sepharose gel and compared with the antibodies purified with protein A and protein G affinity chromatography. The IgG preparations from the histidine affinity column had a much higher catalytic activity in hydrolyzing the peptide substrate Pro-Phe-Arg-methylcoumarinamide compared to the antibodies obtained by the conventional protein A/G method. This preservation of catalytic activity is attributed to the gentle buffer conditions used in the histidine ligand method that allowed the integrity of three-dimensional structure of purified catalytic antibodies. Thus, histidine affinity offer a superior method for isolating autoimmune catalytic antibodies.
Nedonchelle
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Laboratoire d'Interactions Moleculaires et de Technologie des Séparations (LIMTechS), Unité CNRS UPRES A 6022, Centre de Recherche de Royallieu, Compiègne, France.
Pitiot
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Vijayalakshmi
M A
MA
D017151:Antibodies, Catalytic / Q000097:blood / Q000302:isolation & purification* / Q000378:metabolism; D001323:Autoantibodies / Q000097:blood / Q000302:isolation & purification* / Q000378:metabolism; D002846:Chromatography, Affinity / Q000379:methods*; D006639:Histidine; D006801:Humans; D006868:Hydrolysis; D007074:Immunoglobulin G / Q000097:blood / Q000302:isolation & purification / Q000378:metabolism; D066298:In Vitro Techniques; D008024:Ligands; D008180:Lupus Erythematosus, Systemic / Q000276:immunology; D009419:Nerve Tissue Proteins; D009842:Oligopeptides / Q000378:metabolism; D013205:Staphylococcal Protein A
D017151:Antibodies, Catalytic; D001323:Autoantibodies; C029152:G-substrate; D007074:Immunoglobulin G; D008024:Ligands; D009419:Nerve Tissue Proteins; D009842:Oligopeptides; D013205:Staphylococcal Protein A; D006639:Histidine; C025273:prolyl-phenylalanyl-arginine-4-methylcoumaryl-7-amide