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S M Deev

Publications and source records attributed to S M Deev.

At least 19 recordsLinked to original sources
PubMed1991

[Empirical construction of the evolutionary divergence tree of VH-genes of immunoglobulins in mouse family 36-60].

Formerly unknown variable gene of the heavy chain of immunoglobulin (VHTF) was studied in our preceding works in detail. In this paper, we present the results of defining the structure of N-terminus of the heavy chain which confirm the presence of unusual first codon corresponding to threonine. The structure of the VHTF gene is being compared with that of other genes of the 36-60 family, topology of the tree of their evolutionary divergence is being established with the help of empirical algorithm. Its advantages, as compared with the conventional methods, are under discussion.

Animals↗
PubMed1990

[A new expressible VH-gene of the 36-40 family participates in the biosynthesis of antibodies against swine transferrin].

The expressible allelic variant of immunoglobulin heavy chain genes locus from hybridoma PTF-02 is localized in the 17 kb Bgl II fragment of DNA. The structure of VDJh 3 exon and flanking regions was investigated. The 1101 bp fragment was sequenced. It contains transcription regulatory sequences (cd and TTAAAT-box), the leader exon with internal intron (L), the Vh exon, D and Jh 3 segments. The Vh exon has unusual for immunoglobulin genes the first codon-threonine codon. It was shown that expressible in hybridoma PTF-02 Vh gene (Vh TF) represents and earlier unknown gene of the mouse 36-60 Vh family. This conclusion was made after the comparison of Vh TF exon structure and its 5'-region physical map with the other genes of the 36-60 family.

Amino Acid Sequence↗
PubMed1988

[Characteristics of the effect of Ca/Mg-dependent endonuclease from cell nuclei of human lymphocytes on J kappa-cluster of immunoglobulin genes].

The cleavage of the murine J kappa-segments by purified Ca/Mg-dependent endonuclease was studied. The enzyme was shown to cleave specifically the J kappa-segments introducing the double strand breaks in 5'-regions of the J kappa-genes. The number of thus generated discreet fragments does not change during incubation. Specificity of the enzyme is not changed by substitution of Ca/Mg for Mn. The enzyme activity is inhibited by 0.1 M NaCl as well as its specificity. Comparison of the enzyme with the known nuclease activities that cleave specifically the J kappa-segments demonstrated that the activities of the latter are determined by Ca/Mg-dependent endonuclease. The results suggest the possible participation of Ca/Mg-dependent endonuclease in Ig-genes recombination.

Animals↗
PubMed1987

Reciprocal recombination products of VK-JK joining reactions in human lymphoid cell lines.

The recombination process that joins a VK to a JK segment of an immunoglobulin gene generates a second, reciprocal recombination product called f fragment. In this second product the regions flanking the VK and JK segments in the germline are joined in a head to head fashion. We now analysed f fragments in the human lymphoid cell lines Daudi, JI and IARC/BL41. All three f fragments contain JK1 flanks; the VK derived moiety of f Daudi and f41 could be traced back to known germline VK genes. There is a precise head to head joining of the heptanucleotide signal sequences in f Daudi and fJI while in f41 six nucleotides are present between the signal sequences. In contrast to the VK-JK recombination products, the f fragments were found to lack somatic mutations. The structures of the f fragments are discussed in the context of the VK-JK rearrangement mechanism.

Alleles↗
PubMed1986

[Regulator sequences in the kappa-chain gene expressed in the hybridoma PTF.02].

Two kappa genes, one (5.7 kb) from parental myeloma cells that were used for fusion, and the other (7.5 kb) from lymphocytes have been detected in the genome of PTF.02 hybridoma. Functionally important regions of the second gene were sequenced. In the 5'-region, positions and nucleotide sequences of the L-fragment, TAATA- and CAAT-boxes were established. Deca (dc)- and pentadecanucleotide (pd) sequences, obligatory for effective transcription of Kappa genes, were localized at the distance of 91 and 118 bp from the ATG initiating codon of the leader sequence. Together with a true pd, a shadow pd sequence was localized. This sequence was overlapped with the first sequence and shifted by one helical twist. The structure of the region of the enhancer localization was established. Comparison of this sequence with known consensus of the papova virus enhancer allows us to suggest the following structure of the enhancer core for kappa chains: TGTGGCTAA... 10 bp... TGTGGTTA. In the kappa gene under study, the variable fragment is linked to the J5 segment. In the latter, a point mutation C----G was found, to which a conservative substitution Ala - Gly in a hypervariable region of the chi-chain should correspond. Somatic mutations were also observed within the intron region adjacent to the J5 segment.

Animals↗
PubMed1982

[Construction and selection of bacterial clones hybridized with the mRNA of a light-chain immunoglobulin].

The copy-DNA was synthesized on the mRNA fraction which was isolated from MOPC 21 mice myeloma by reverse transcription. This copy was completed with another chain without adding the exogenous primer, with the aid of the Klenov fragment of DNA polymerase I. After treatment of the double-stranded pin DNA with endonuclease S1, the poly(dA) sequences were built up using terminal deoxynucleotidyl transferase. The building up of the poly(dT) sequences at the DNA 3'-termini of pBR322 plasmid previously treated with BamHI was performed in a similar way. The average length of connecting polynucleotide sequences was 50 nucleotides. After annealing and transformation of the cells with Escherichia coli hybrid plasmids, selection of clones was made for ApRTcS phenotype. Further, we applied a stepwise selection of clones by means of increasing the specificity: colony hybridization, quantitative hybridization of the excess of plasmid DNA with (32P)-cDNA and hybridization of plasmid DNAs with mRNA units which were transferred from electrophoregrams to the diazo-papers. As a result, bacterial clones were selected which contained gene fragments showing the ability to hybridize with the RNA fraction characterized by mRNA mobility of the light-chain immunoglobulin G.

Animals↗
PubMed1982

[Analysis and properties of a bacterial clone containing a gene fragment of an immunoglobulin L chain].

Restriction analysis of hybrid plasmids from a number of clones allowed to screen out the plasmid p8-1. A detailed analysis of this plasmid and that of the inserted DNA was made using HhaII, AluI, Sau96I, Sau3A, MspI and BspI restriction endonucleases. The results demonstrated that the inserted DNA corresponds to the 3'-nontranslated region and to a portion of the C fragment of the light-chain immunoglobulin gene. Establishing the partial structure of the recombinant plasmid exhibited a complete coincidence of the inserted DNA 3'-terminus and the primary structure of the light-chain mRNA synthesized by MOPC-21 myeloma cells. This sequence corresponds to the amino acid sequence in the light-chain constant region (207-214 residues). The analysis of p8-1 plasmid showed that nucleotides of pBR322 plasmid at positions from 376 to 618 were deleted. The deletion might be predetermined by the plasmid property to be amplified more readily than all the other hybrid plasmids which were not deleted.

Animals↗
PubMedDate not supplied

[Immunoglobulin genes in lymphoid cells and regulation of their transcription].

The hybridoma genomes contain polyploid sets of immunoglobulin genes. We have shown, that the hybridoma PTF-02 genome contains three genes of heavy chains and two genes of light chains. The genes responsible for antibody synthesis were cloned and their structure were determined. Investigation of the kappa gene transcription and its fragments which contain regulatory sequences revealed a nuclear factor. The latter interacts with the octanucleotide localized at the promoter region of the kappa gene. The purified factor activates the transcription of the kappa gene in a heterologous cell-free system. Together with the tissue-specific factor there is also an universal factor interacting with the octanucleotide sequence. We have shown an additional factor in lymphoid cells interact with the protein which binds to the octanucleotide sequence. We have shown an additional factor in lymphoid cells interacting with the protein which binds to the octanucleotide sequence. As a result, there is a family of factors which interact with ATTTGCAT sequence. One major factor (m.w. 60 +/- 2 kDa) is an obligatory component for the initiation of immunoglobulin genes transcription.

Animals↗
PubMedDate not supplied

[Immunoglobulin kappa-genes. Cloning, hybridization and structural analysis].

The principal problems in molecular and genetic immunology to be resolved are the structure of Ig-genes and the regulation of their expression. The isolation of mRNA for light and heavy Ig-chains would be a first step along this line. A combination of two approaches may be the best strategy in mRNA preparation. Affinity purification allows one to obtain pure mRNA in a one-step procedure whereas immunoprecipitation makes it possible to prepare mRNAs for both heavy and light immunoglobulin chains in considerable amounts. In the series of experiments, conditions for synthesis of long and short cDNA chains were elaborated and the clone containing the fragment of kappa-chain gene was isolated and characterized. This clone was used as a probe for hybridization with genomic DNA from myeloma and hybridoma cells. It was shown that both allelic genes on homologous chromosomes were rearranged in myeloma MOPC21 cells. The original cell line of hybridoma PTF02 contains the embryonic gene as well as the differentiated genes. However, only differentiated genes can be detected in a similar experiment conducted with the same hybridoma after passage on mice. In conclusion, the coordination of homologous and heterologous chromosome expression in B cells in discussed in terms of the feed-back control.

Amino Acid Sequence↗
PubMedDate not supplied

[Synthesis of double-stranded DNA on light immunoglobulin chain matrix RNA].

cDNA was synthesized on Ig kappa-L-chain mRNA isolated from mouse plasmocytoma MOPC 21 using avial myeloblastosis RNA-dependent DNA polymerase. The cDNA was used as a matrix for second DNA chain synthesis using 5'-exonuclease free DNA-polymerase I (Klenoff fragment). Hairpin DNA having double length of initial cDNA was obtained without added exogenous primer. SI nuclease treatment of the hairpin DNA results in reducing the DNA length twice, as shown by electrophoresis in denaturing conditions. The fact, that S1 nuclease resistance of the hairpin DNA is 75% shows high complementarity between first and second DNA chain. The product length obtained after S1 nuclease treatment was shown to be heterogenous. The maximal length of the product, reaching at least 900 base pairs, is near to be initial mRNA length (without the poly(A)-fragment).

Animals↗
PubMedDate not supplied

[Group-selective immunoanalysis].

The new solid-phase immunoassay technique has been applied for antidigoxin monoclonal antibodies (Mabs) detection. At the first assay step, antidigoxin Mabs (IgG1, Kaff = 9.2 x 10(9) M-1) bound with specially prepared immunosorbent: microtiter plates with adsorbed digoxin-human serum albumin conjugate, in which free amino groups were blocked by glutaraldehyde (the microtiter plates with immobilized conjugate were treated for 3h by 2.5% glutaraldehyde at 45 degrees C, this had no significant effect on the immunological properties of immunosorbents). After antigen-antibody reaction and solid phase separation, free amino groups were located only at the antidigoxin Mabs bound to GSI immunosorbent. At the second assay step, N-hydroxysuccinimide-biotin ester solution was added to bind with the free amino groups. After separation at the third assay step, biotin labels were detected by the streptavidin-peroxidase conjugate. The method does not require second labeled antibodies and may be used for anti-hapten screening.

Antibodies, Monoclonal↗