Metal dependent hydrolysis of β-casein by sIgA antibodies from human milk.
We present the first evidence that electrophoretically and immunologically homogeneous sIgAs purified from milk of healthy human mothers by chromatography on Protein A-Sepharose and FPLC gel filtration contain intrinsically bound metal ions (Ca  >  Mg ≥ Al > Fe approximately  Zn ≥ Ni ≥ Cu ≥ Mn), the removal of which by a dialysis against ethylenediamine tetraacetic acid (EDTA) leads to a significant decrease in the β-casein-hydrolyzing activity of these antibodies (Abs). An affinity chromatography of total sIgAs on benzamidine-Sepharose interacting with canonical serine proteases separates a small metalloprotease sIgA fraction (6.8 ± 2.4%) from the main part of these Abs with a serine protease-like β-casein-hydrolyzing activity. The relative activity of this metalloprotease sIgA fraction containing intrinsically bound metal ions increases ∼1.2-1.9-fold after addition of external metal ions (Mg(2+) > Fe(2+) > Cu(2+) ≥ Ca(2+) ≥ Mn(2+)) but decreases by 85 ± 7% after the removal of the intrinsically bound metals. The metalloprotease sIgA fraction free of intrinsic metal ions demonstrates a high β-casein-hydrolyzing activity in the presence of individual external metal ions (Fe(2+) > Ca(2+) > Co(2+) ≥ Ni(2+)) and especially several combinations of metals: Co(2+) + Ca(2+) < Mg(2+) + Ca(2+) < Ca(2+) + Zn(2+) < Fe(2+) + Zn(2+) < Fe(2+) + Co(2+) < Fe(2+) + Ca(2+). The patterns of hydrolysis of a 22-mer oligopeptide corresponding to one of sIgA-dependent specific cleavage sites in β-casein depend significantly on the metal used. Metal-dependent sIgAs demonstrate an extreme diversity in their affinity for casein-Sepharose and chelating Sepharose, and interact with Sepharoses bearing immobilized monoclonal mouse IgGs against λ- and κ-type light chains of human Abs. Possible ways of the production of metalloprotease abzymes (Abz) by human immune system are discussed.
Institute of Chemical Biology and Fundamental Medicine, Siberian Division of Russian Academy of Sciences, 8, Lavrentiev Ave., Novosibirsk 630090, Russia.
Elena S
ES
Odintsova
Nataliya P
NP
Zaksas
Valentina N
VN
Buneva
Georgy A
GA
Nevinsky
D000328:Adult; D017151:Antibodies, Catalytic / Q000302:isolation & purification / Q000378:metabolism; D002364:Caseins / Q000378:metabolism*; D005260:Female; D006801:Humans; D006868:Hydrolysis; D007071:Immunoglobulin A, Secretory / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D045726:Metalloproteases / Q000302:isolation & purification / Q000378:metabolism; D008670:Metals / Q000378:metabolism*; D008895:Milk, Human / Q000276:immunology / Q000378:metabolism; D055815:Young Adult
D017151:Antibodies, Catalytic; D002364:Caseins; D007071:Immunoglobulin A, Secretory; D008670:Metals; D045726:Metalloproteases