title: Systemic lupus erythematosus: molecular cloning and analysis of recombinant DNase monoclonal κ light chain NGK-1.
abstract: Because DNase antibodies are cytotoxic, enter the nucleus and cause DNA fragmentation inducing cell death by apoptosis, they can play an important role in the pathogenesis of different autoimmune pathologies and especially systemic lupus erythematosus (SLE). The interesting goal of catalytic antibodies research is not only to study a possible biological role of such antibodies, but also to develop in future new human and animal therapies that use the advantages offered by abzymes. An immunoglobulin κ light chain library from SLE patients was cloned into a phagemid vector. Phage particles displaying recombinant monoclonal antibody light chains (MLChs) capable of binding DNA were isolated by affinity chromatography on DNA-cellulose. Sixteen of the 46 MLChs efficiently hydrolyzed DNA; one MLCh (approximately 27-28kDa) was expressed in Escherichia coli and purified by metal chelating and gel filtration. MLCh NGK-1 was electrophoretically homogeneous and demonstrated a positive answer with mouse IgGs against light chains of human antibodies after western blotting. SDS-PAGE in a gel containing DNA demonstrated that the MLCh hydrolyzes DNA and is not contaminated by canonical DNases. The DNase MLCh was activated by several metal ions. The protein sequence of the DNase MLCh has homology with mammalian DNases I and shares with them several identical or similar (with the same side chain functionality) important amino acid residues, which are necessary for DNA hydrolysis and binding of Mg(2+) and Ca(2+) ions. The affinity of DNA for this first example of a MLCh (K(M) = 0.3 microM) was 150- to 200-fold higher than for human DNase I.
authors.affiliation: Institute of Chemical Biology and Fundamental Medicine, Siberian Division of Russian Academy of Sciences, Novosibirsk 630090, Russia.
authors.forename: Irina A
authors.initials: IA
authors.lastname: Kostrikina
authors.affiliation: Institute of Chemical Biology and Fundamental Medicine, Siberian Division of Russian Academy of Sciences, Novosibirsk 630090, Russia.
authors.forename: Elena S
authors.initials: ES
authors.lastname: Odintsova
authors.affiliation: Institute of Chemical Biology and Fundamental Medicine, Siberian Division of Russian Academy of Sciences, Novosibirsk 630090, Russia Novosibirsk State University, Novosibirsk 630090, Russia.
authors.forename: Valentina N
authors.initials: VN
authors.lastname: Buneva
authors.affiliation: Institute of Chemical Biology and Fundamental Medicine, Siberian Division of Russian Academy of Sciences, Novosibirsk 630090, Russia Novosibirsk State University, Novosibirsk 630090, Russia nevinsky@niboch.nsc.ru.
authors.forename: Georgy A
authors.initials: GA
authors.lastname: Nevinsky
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D017151:Antibodies, Catalytic; D001483:Base Sequence; D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism; D003851:Deoxyribonucleases / Q000276:immunology* / Q000378:metabolism; D004789:Enzyme Activation; D006801:Humans; D006868:Hydrolysis; D007145:Immunoglobulin kappa-Chains / Q000737:chemistry / Q000235:genetics* / Q000276:immunology*; D007700:Kinetics; D008180:Lupus Erythematosus, Systemic / Q000235:genetics* / Q000276:immunology* / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D016415:Sequence Alignment
chemical_list: D017151:Antibodies, Catalytic; D007145:Immunoglobulin kappa-Chains; D011994:Recombinant Proteins; D004247:DNA; D003851:Deoxyribonucleases
keywords: DNase activity; homology with mammalian DNases I; recombinant monoclonal light chain; systemic lupus erythematosus