title: Systemic lupus erythematosus: molecular cloning and analysis of recombinant monoclonal kappa light chain NGTA2-Me-pro-ChTr possessing two different activities-trypsin-like and metalloprotease.
abstract: Polyclonal antibodies hydrolyzing myelin basic protein (MBP) can play an important role in the pathogenesis of multiple sclerosis and systemic lupus erythematosus (SLE). An immunoglobulin light chain phagemid library derived from peripheral blood lymphocytes of patients with SLE was used. The small pools of phage particles displaying light chains with different affinity for MBP were isolated by affinity chromatography on MBP-Sepharose. The fraction eluted with 0.5M NaCl was used for preparation of individual monoclonal light chains (MLChs, 26-27kDa). The clones were expressed in Escherichia coli in a soluble form; MLChs were purified by metal-chelating chromatography followed by gel filtration. In mammalians, there are serine proteases and metalloproteases. These and many other enzymes usually have only one active site and catalyze only one chemical reaction. In contrast to canonical proteases, one MLCh (NGTA2-Me-pro-ChTr) efficiently hydrolyzed MBP (but not other proteins) and four different oligopeptides corresponding to four immunodominant sequences containing cleavage sites of MBP. The proteolytic activity of MLCh was efficiently inhibited only by specific inhibitors of serine-like (phenylmethanesulfonylfluoride, PMSF) and metalloproteases (EDTA). It was shown that MLCh possess independent serine-like and metal-dependent activities. The principal existence of monoclonal antibodies with two different proteolytic activities is unexpected but very important for the further understanding of at present unknown biological functions of human antibodies.
authors.affiliation: Institute of Chemical Biology and Fundamental Medicine, Siberian Division of Russian Academy of Sciences, Novosibirsk 630090, Russia.
authors.forename: Anna M
authors.initials: AM
authors.lastname: Timofeeva
authors.affiliation: Institute of Cytology and Genetics, Siberian Division of Russian Academy of Sciences, Novosibirsk 630090, Russia.
authors.forename: Nikita V
authors.initials: NV
authors.lastname: Ivanisenko
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: D017151:Antibodies, Catalytic / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D004492:Edetic Acid / Q000737:chemistry; D004926:Escherichia coli / Q000235:genetics*; D006801:Humans; D016056:Immunodominant Epitopes / Q000276:immunology / Q000378:metabolism*; D007145:Immunoglobulin kappa-Chains / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008180:Lupus Erythematosus, Systemic / Q000201:enzymology / Q000276:immunology*; D045726:Metalloproteases / Q000737:chemistry / Q000378:metabolism*; D004676:Myelin Basic Protein / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D019151:Peptide Library; D010664:Phenylmethylsulfonyl Fluoride / Q000737:chemistry; D057057:Serine Proteases / Q000737:chemistry / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D017151:Antibodies, Catalytic; D016056:Immunodominant Epitopes; D007145:Immunoglobulin kappa-Chains; D004676:Myelin Basic Protein; D010446:Peptide Fragments; D019151:Peptide Library; D010664:Phenylmethylsulfonyl Fluoride; D004492:Edetic Acid; D045726:Metalloproteases; D057057:Serine Proteases
keywords: hydrolysis of myelin basic protein; recombinant monoclonal light chain; similar amino acids of active center with serine-like and metalloproteases; systemic lupus erythematosus