DNA-specific antiidiotypic antibodies in the sera of patients with autoimmune diseases.

Blood sera of patients with autoimmune diseases scleroderma (Scl), systemic lupus erythematosus (SLE), and rheumatoid arthritis (RA) have been shown to yield a specific immune response to topoisomerase I, the product of expression of a cDNA fragment cloned into lambda gt11 and monoclonal antibodies (MAB) to the enzyme. The 'topoisomerase test' is not absolutely specific for Scl. The stable positive response of autoimmune sera to anti-topoisomerase monoclonal antibodies has a specific character and is associated with the interaction of the Fab fragment of MAB with the IgG fraction of autoimmune serum. The response observed indicates the induction of anti-idiotypic antibodies against topoisomerase. The anti-idiotype, isolated by HPLC and affinity chromatography demonstrated the following functional activities: (i) the immunological reaction against DNA; (ii) high-affinity DNA-binding with topoisomerase-specific consensus; (iii) ability to compete with the native enzyme for binding with DNA and MAB to topoisomerase; (iv) immunological reaction against MAB to topoisomerase.

[Interactions of sera from patients with autoimmune diseases with cDNA expressed fragment of topoisomerase I and monoclonal antibodies against enzyme]. / Vzaimodeistvie syvorotok krovi bol'nykh autoimmunnymi zabolevaniiami s ékspressirovannym fragmentom kDNK topoizomerazy I i monoklonal'nymi antitelami k fermentu.

The interaction of sera from 34 patients with different autoimmune diseases with the expressed fusion protein cloned in lambda gt11 vector (topoisomerase I--beta galactosidase) and monoclonal antibodies against enzyme was studied. It was demonstrated that 100% of Scl cases possessed positive activity against fusion protein. It was shown that this test is not absolutely specific for Scl, i. e. 57.1% of Sle and 84.6% of RA demonstrated positive activity against "topoisomerase test". Autoimmune sera had the positive activity against monoclonal antibodies for topoisomerase I. This activity was shown to be due to the presence of antiidiotypic antibodies against topoisomerase in the autoimmune sera.

[Topoisomerase I from human placenta. Functional activity of products of expression of cloned cDNA fragments]. / Topoizomeraza I iz platsenty cheloveka. Funktsional'naia aktivnost' produktov ékspressii klinirovannykh fragmentov kDNK.

Immunoscreening of the human placenta cDNA-library in the expression vector lambda gt11 using non-isotope detection based on the avidin-biotin system allowed to identify a number of clones encoding human topoisomerase I. The fusion protein from an extract of Escherichia coli cells infected with the recombinant phage lambda gt11 interacts with the monoclonal antibody raised against topoisomerase I from calf thymus; the dissociation constant being 5.7.10(-8) M. The restricted DNA fragments coding for the topoisomerase polypeptide in the composition of the fusion protein were recloned, and expression in the pEX vector was obtained. The functional analysis of the expression products has enabled localization of the epitope of binding the monoclonal antibody. It was demonstrated that the identified fusion protein can be applied for diagnosis of autoimmune diseases.

[Specific splitting of the alpha-A-globin gene by molecular forms of DNA-topoisomerase type I from calf thymus]. / Spetsificheskoe rasshcheplenie alpha-A-globinovogo gena molekuliarnymi formami DNK-topoizomeraz I tipa iz timusa telenka.

The specificity of splitting of the cloned alpha A-globin gene from chicken erythrocytes induced by three topoisomerases I differing in molecular masses was demonstrated. The localization and relative number of topoisomerase breaks in the alpha A-globin gene vary in different topoisomerase I forms.

[Effect of camptothecin on the DNA-relaxing and DNA-cleavage activity of calf thymus topoisomerase I]. / Vliianie kamptotetsina na DNK-relaksiruiushchuiu i DNK-rasshchepliaiushchuiu aktivnosti topoizomerazy I iz timusa telenka.

Nucleotide sequences that are cleaved by calf thymus type I topoisomerase have been determined using cloned human Ha-ras and p53 genes. Localization and relative frequency of single-strand cleavages within these sequences were observed to change in the presence of the cytotoxic alkaloid camptothecin.

[DNA-topoisomerases of animal cells]. / DNK-topoizomerazy zhivotnykh kletok.

The properties and functions of the DNA topoisomerases are reviewed on the basis of the literature and the author's own data. The techniques of isolation and characterization of the covalent complexes of the topoisomerases with DNA are described.

[Isolation and properties of endodeoxyribonuclease from the ovicells of the loach]. / Vydelenie i svoistva éndodezoksiribonukleazy iz iaitsekletok v'iuna.

A new DNAase was isolated from unfertilized loach eggs. The enzyme is activated by bivalent metal ions (Mg2+, Ca2+, Mn2+) and is inhibited by Na+ and N-ethylmaleimide. The maximal activity of DNAase lies within the pH range of 9.0-9.5. The enzyme is thermostable and predominantly hydrolyzes the two-chain DNA by producing one-chain ruptures. The DNAase activity during the purification procedure and enzymatic analysis was determined by a highly sensitive method based on registration of superhelical plasmid or phage DNA conversion into open circular or linear molecules.

[Changes in the representation of repeating DNA sequences in nuclear RNAs of early loach embryos]. / Izmeneniia predstavlennosti povtoriaiushchikhsia posledovatel'nostei DNK v iadernykh RNK rannikh émbrionov v'iuna.

The hybridization kinetics of nuclear RNAs of loach embryos labelled with [3H]uridine for 1 hour with DNA excess shows that during embryogenesis (from the blastula to the gastrula stage) the number of newly formed RNA molecules transcribed from repeating DNA sequences in considerably reduced. This occurs both in the RNA fraction extracted from the nuclei with phenol pH 7.7 and having a low sedimentation coefficient and a low degree of polyadenylation, and in the RNA fraction extracted with phenol pH 9.0 having a higher sedimentation coefficient and a higher degree of polyadenylation.