Browsing by Author "Sabarinath, PS"

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    Dual Specificity of Human Plasma Lactose-Binding Immunoglobulin to Anomers of Terminal Galactose Enables Recognition of Desialylated Lipoprotein(a) and Xenoantigens
    (SCANDINAVIAN JOURNAL OF IMMUNOLOGY, 2014) Sabarinath, PS; Chacko, BK; Appukuttan, PS
    Human plasma lactose-binding immunoglobulin (LIg) isolated by affinity chromatography on lactose-Sepharose was largely IgG with significant IgA and IgM contents. LIg-mediated agglutination of desialylated human RBC was inhibited equally by the alpha- and beta-anomers of methyl galactoside. Recognition of either the terminal alpha-galactose (TAG)-containing glycans of bovine thyroglobulin or the N-acetyl lactosamine (LacNAc)-terminating glycans of asialofetuin by LIg was inhibitable nearly as much by the alpha-galactoside melibiose as by the beta-galactoside lactose. Melibiose covalently conjugated to protein and coated on polystyrene wells captured several times more LIg molecules than its lactose analogue. LIg binding to bovine thyroglobulin or rabbit RBC membrane proteins, both bearing TAG was substantially reduced by prior treatment of the proteins with alpha-galactosidase to remove TAG though enzyme-treated glycans contained newly exposed LacNAc moieties. Desialylated O-linked oligosaccharides, however, were no ligand for LIg. Unlike LDL, plasma lipoprotein(a) [Lp(a)] coated on polystyrene well and desialylated by neuraminidase was recognized by LIg through terminal LacNAc moieties exposed by the enzyme on its apo(a) subunit. Further, same amount of added fluorescence-labelled LIg formed significantly more immune complex with Lp(a) in high Lp(a) plasma than in low Lp(a) plasma. Results suggest (1) possibility of a role for LIg in combating non-primate molecules and cells bearing TAG moiety and (2) a mechanism for Lp(a)-mediated vascular injury as diabetes, infections and inflammations induce greater release of neuraminidase into circulation.
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    Immunopathology of desialylation: human plasma lipoprotein(a) and circulating anti-carbohydrate antibodies form immune complexes that recognize host cells
    (MOLECULAR AND CELLULAR BIOCHEMISTRY, 2015) Sabarinath, PS; Appukuttan, PS
    Human plasma lipoprotein(a) [Lp(a)], the dominant lipoprotein in atherosclerotic plaques, contains an apo(a) subunit of variable size linked to the apoB subunit of a low-density lipoprotein (LDL) molecule. Circulating lipoprotein immune complexes (ICs) assayed by ELISA using microplate-coated anti-apo(a) or anti-apoB antibody for capture and peroxidase-labelled anti-human immunoglobulins as probe consisted mostly of Lp(a) despite several-fold excess of LDL over Lp(a) in plasma. Microplate coating of plasma lipoprotein IC and probing with antibodies to apo(a) and apoB also revealed negligible presence of LDL compared to Lp(a). Peanut agglutinin specific to desialylated O-glycans bound significantly more to Lp(a) recovered after urea dissociation of IC than to free Lp(a). Plasma lipoproteins separated by ultracentrifugation and desialylated by neuraminidase formed IC with naturally occurring antibodies in normal plasma. These de novo ICs agglutinated desialylated but not normal human RBC in proportion to the polyagglutinin antibody titre of plasma used, suggesting availability of multiple unoccupied binding sites on the participating antibodies even after IC formation. Agglutination was inhibitable by galactosides and decreased 4-8 fold if precursor lipoprotein was selectively depleted of Lp(a), showing agglutinating ICs were contributed mainly by desialylated Lp(a) and galactose-specific antibodies. IC was 2 fold more agglutinating if lipoproteins used contained smaller rather than larger Lp(a) molecules of the same number. Small size/high plasma concentration Lp(a) phenotype and neuraminidase-releasing diseases including diabetes are risk factors for vascular disorders. Results suggest a possible route of Lp(a) attachment to vascular cells that offer terminal galactose on surface glycans following desialylation.
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    Plasma anti-alpha-galactoside antibody binds to serine- and threonine-rich peptide sequence of apo(a) subunit in Lp(a)
    (GLYCOCONJUGATE JOURNAL, 2014) Geetha, M; Kalaivani, V; Sabarinath, PS; Appukuttan, PS
    Lipoprotein(a) immune complexes [Lp(a) IC] of varying particle density obtained by ultracentrifugation of plasma from normal healthy donors were markedly dominated by IgG. Lp(a) and immunoglobulins were liberated from plasma Lp(a) IC by treatment with melibiose, a sugar specific for circulating anti-alpha-galactoside antibody (anti-Gal). Upon incubation with plasma lipoprotein fraction anti-Gal but not the alpha-glucoside-specific antibody from human plasma formed de novo IC with Lp(a). Binding of Lp(a) sugar-reversibly enhanced the fluorescence of FITC-labeled anti-Gal as did binding of alpha-galactoside-containing glycoproteins. This effect apparently due to conformational shift in the Fc region of the antibody was also produced by apo(a) subunit separated from Lp(a) and de-O-glycosylated apo(a) but not by any other plasma lipoproteins or by Lp(a) pre-incubated with the O-glycan-specific lectin jacalin. O-Glycans and their terminal sialic acid moieties in apo(a) of circulating Lp(a)-anti-Gal IC, in contrast to those in pure Lp(a), were inaccessible to jacalin and anion exchange resin, respectively. Unlike other plasma lipoproteins, Lp(a) inhibited Griffonia simplicifolia isolectin B4 which also accommodates serine- and threonine-rich peptide sequence (STPS) as surrogate ligand to alpha-galactosides at its binding site. Results suggest that anti-Gal recognizes STPS in the O-glycan-rich regions of apo(a) subunit in Lp(a) which contains no alpha-linked galactose.