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Biomedical subjects

V M Lipkin

Publications and source records attributed to V M Lipkin.

At least 37 records · Page 2Linked to original sources
PubMed2000

Leptin and apoptosis inhibitor soluble Fas antigen in the serum of patients with osteosarcoma and neuroectodermal bone tumors.

Serum contents of leptin and soluble Fas antigen were measured in 28 patients with osteosarcoma, 7 with neuroectodermal bone tumors, and 17 healthy subjects. The incidence and levels of soluble Fas antigen in patients with osteosarcoma and neuroectodermal bone tumors were higher than in healthy subjects and did not depend on sex and age in both healthy subjects and patients. Serum concentration of leptin in women was higher than in men (both in healthy controls and patients) and was lower in healthy subjects compared to patients with osteosarcoma and neuroectodermal bone tumors. A trend to a negative correlation between the concentrations of leptin and soluble Fas antigen in female patients with osteosarcoma and male patients with neuroectodermal bone tumors was revealed. The role of leptin and soluble Fas antigen in the pathogenesis of primary bone tumors is discussed.

Adolescent↗
PubMed2000

Influence of synthetic peptide corresponding to the ACTH-like sequence of human immunoglobulin G1 on proliferation of lymphoblastoid cells.

Influence of the ACTH-like peptide H-Val-Lys-Lys-Pro-Gly-Ser-Ser-Val-Lys-Val-OH corresponding to the sequence 11-20 of the variable part of human immunoglobulin G1 (IgG1) heavy chain on growth of MT-4 human T-lymphoblastoid cell line was investigated. It was found that the ACTH-like peptide at concentration range 10(-11) -10(-7) M inhibits the proliferation of MT-4 cells. Labeled ACTH 'address segment' [(125)I]ACTH (13-24) was used to establish that MT-4 cells express specific receptors for ACTH (K(d) = 97 pM). The ACTH-like peptide and human ACTH (but not IgG1 heavy chain) were shown to compete with [(125)I]ACTH (13-24) for binding to these receptors (K(i1) = 0.38 nM and K(i2) = 0.34 nM).

Adrenocorticotropic Hormone↗
PubMed2000

Effects of mutations in the calcium-binding sites of recoverin on its calcium affinity: evidence for successive filling of the calcium binding sites.

A molecule of the photoreceptor Ca(2+)-binding protein recoverin contains four potential EF-hand Ca(2+)-binding sites, of which only two, the second and the third, are capable of binding calcium ions. We have studied the effects of substitutions in the second, third and fourth EF-hand sites of recoverin on its Ca(2+)-binding properties and some other characteristics, using intrinsic fluorescence, circular dichroism spectroscopy and differential scanning microcalorimetry. The interaction of the two operating binding sites of wild-type recoverin with calcium increases the protein's thermal stability, but makes the environment around the tryptophan residues more flexible. The amino acid substitution in the EF-hand 3 (E121Q) totally abolishes the high calcium affinity of recoverin, while the mutation in the EF-hand 2 (E85Q) causes only a moderate decrease in calcium binding. Based on this evidence, we suggest that the binding of calcium ions to recoverin is a sequential process with the EF-hand 3 being filled first. Estimation of Ca(2+)-binding constants according to the sequential binding scheme gave the values 3.7 x 10(6) and 3.1 x 10(5) M(-1) for third and second EF-hands, respectively. The substitutions in the EF-hand 2 or 3 (or in both the sites simultaneously) do not disturb significantly either tertiary or secondary structure of the apo-protein. Amino acid substitutions, which have been designed to restore the calcium affinity of the EF-hand 4 (G160D, K161E, K162N, D165G and K166Q), increase the calcium capacity and affinity of recoverin but also perturb the protein structure and decrease the thermostability of its apo-form.

Calcium↗
PubMed2000

Structure and mechanisms of function of visual system proteins.

Recent progress in understanding visual signal transduction in retinal cells is summarized. The roles of particular proteins in activation, amplification and termination of the photoresponse are described. Detailed information on the structure and function of the photoreceptor protein rhodopsin is presented. The latest data on visual pigment sequences, rhodopsin mutations in the autosomal-dominant retinitis pigmentosa, and the results of site-directed mutagenesis of the rhodopsin molecule are summarized.

Amino Acid Sequence↗
PubMed2000

[Hormone-like activity of a synthetic decapeptide with the adrenocorticotropin-like sequence of human immunoglobulin G1].

The antiproliferative and immunosuppressive in vitro effects of immunocortin, a synthetic adrenocorticotropin-like (ACTH-like) decapeptide H-Val-Lys-Lys-Pro-Gly-Ser-Ser-Val-Lys-Val-OH, whose sequence corresponds to segment 11-20 of the variable part of the human IgG1 heavy chain, were studied. At concentrations of 10(-11)-10(-7) M, immunocortin was found to inhibit the growth of the human MT-4 T-lymphoblastoid cell line, to suppress the blast transformation of thymocytes, and to decrease the spontaneous mobility of peritoneal macrophages and their bactericidal action toward the virulent strain Salmonella typhimurium 415. By using a 125I-labeled "addressing" fragment of ACTH ¿[125I]ACTH-(13-24)¿, we showed that MT-4 cells express specific receptors for ACTH (Kd 97 pM). Immunocortin and human ACTH (but not the heavy chain of IgG1) competitively inhibited the binding of [125I]ACTH-(13-24) to these receptors with Ki1 of 0.38 and Ki2 of 0.34 nM, respectively. Specific receptors for ACTH (Kd 5.8 nM) on mouse thymocytes were detected and characterized. The unlabeled immunocortin was shown to complete with labeled ACTH-(13-24) for binding to these receptors (Ki = 1.8 nM) and this binding of immunocortin to receptors on thymocytes activates adenylate cyclase from these cells and increases the intracellular concentration of cAMP.

Adrenocorticotropic Hormone↗
PubMed2000

[Single amino acid substitutions in the Ca2+-binding site of recoverin.II.The unusual behavior of the protein upon the binding of calcium ions].

The structural properties of myristoylated forms of recombinant recoverin of the wild type and of its mutants with damaged second and/or third Ca(2+)-binding sites were studied by fluorimetry and circular dichroism. The interaction of wild-type recoverin with calcium ions was shown to induce unusual structural rearrangements in its molecule. In particular, protein binding with Ca2+ ions results in an increase in the mobility of the environment of Trp residues, in higher hydrophobicity, and in elevated thermal stability (its thermal transition shifts by 15 degrees C to higher temperatures) but has almost no effect on its secondary structure. Similar structural changes induced by Ca2+ are also characteristic of the -EF2 mutant of recoverin whose second Ca(2+)-binding site is modified and cannot bind calcium ions. The structural properties of the -EF3 and -EF2,3 mutants (whose third or simultaneously second and third Ca(2+)-binding sites, respectively, are modified and damaged) are practically indifferent to calcium ions.

Amino Acid Substitution↗
PubMed2000

[Point amino acid substitutions in Ca2+-binding centers of recoverin. III. Mutant with the fourth reconstructed Ca2+-binding site].

Unlike wild type recoverin with only two (the second and the third) functioning Ca(2+)-binding sites out of four potential ones, the +EF4 mutant contains a third active Ca(2+)-binding site. This site was reconstructed from the fourth potential Ca(2+)-binding domain by the introduction of several amino acid substitutions in it by site-directed mutagenesis. The effect of these mutations in the fourth potential Ca(2+)-binding site of myristoylated recoverin on the structural features and conformational stability of the protein was studied by fluorimetry and circular dichroism. The apoform of the resulting mutant (free of Ca2+ ions) was shown to have a higher calcium capacity, significantly lower thermal stability, and noticeably different secondary and tertiary structures as compared with the apoform of wild type recoverin.

Amino Acid Substitution↗
PubMed2000

[The endogenous differentiation factor of the HL-60 cells shows a nuclease activity].

A structural homology between the endogenous differentiation factor of the HL-60 cell line of promyelocyte leukemia (HLDF) and several DNA/RNA-binding and DNA/RNA-hydrolyzing proteins was revealed, and expression of the hldf gene in prokaryotic systems was studied. On the basis of these experiments, the amino acid sequence of an 8-membered fragment of HLDF with potential nuclease activity was identified. The synthetic octapeptide RRWHRLKE was shown to be capable of the cleavage of RNA, linear DNA from phage lambda, and all forms of plasmid DNA. We established that treatment of the HL-60 cell culture with this peptide (10(-6) M) results in an increase in the number of apoptotic cells and suggested that HLDF is involved in processes of apoptosis.

Amino Acid Sequence↗
PubMed2000

Cloning and characterization of the human ribosomal protein S21 gene [.

A full-size functional gene encoding the human ribosomal protein S21 was cloned and characterized. Its nucleotide sequence, exon-intron organization, and transcription initiation site were determined. The gene comprises 1417 bp and is composed of six exons and five introns. Like most known genes of mammalian ribosomal proteins, it lacks the canonical TATA- and CAAT sequences in the promoter region and harbors potential binding sites for transcription factors both upstream and downstream the transcription initiation site. The first intron of the rpS21 gene is located in the 5'-untranslated region. The transcription initiation site is at a 53-bp distance from the ATG codon, and the initiation cytidine is surrounded by polypyrimidine tracts. The 5'-flanking region contains two repeats belonging to the Alu-S and Alu-J families.

Base Sequence↗
PubMed2000

[Deletion mutants of the natriuretic peptide receptor type B: expression of cDNA, purification and characteristics of proteins].

The C type natriuretic peptide (CNP) is a peptide hormone stimulating vasorelaxation and inhibiting cell proliferation. CNP activates the type B natriuretic peptide receptor (NPR-B), known as the guanylate cyclase B membrane enzyme, which results in the cGMP release. To study functional properties of NPR-B, its gene fragments were expressed in methylotrophic yeasts Pichia pastoris. Conditions were found providing for secretion of functionally active recombinant proteins NPR-Bs and NPR-B1 into the cultural medium in a yield of 25 mg/ml culture. Their specific activity was 0.97 and 0.93 mumol cGMP min-1 mg-1 protein, respectively. It was shown that NPR-B belongs to the family of Ser/Thr protein kinases and can be autophosphorylated at the serine residues.

Base Sequence↗
PubMed2000

[Biologically active fragment of the differentiation factor from HL-60 cell line. Identification and properties].

Six-membered peptide fragment TGENHR (HLDF-6) was identified in the HL-60 cell culture of human promyelocyte leukemia treated with retinoic acid when studying the differentiation factor HLDF of this cell line. HLDF-6 retains the ability of the full-size factor to induce the differentiation and arrest the proliferation of the starting HL-60 cells. It was shown that the synthetic peptide HLDF-6 has no specific receptors on the surface of the HL-60 cells but can affect the binding of interleukin IL-1 beta, a cytokine involved in proliferation, to the cell surface. It was found on a model of transplantable NSO myeloma that HLDF-6 has an antitumor activity.

Animals↗
PubMed2000

[Biological role of a neurotrophic factor fragment from pigment epithelium: structure-functional homology with a differentiation factor for the HL-60 cell line].

It was shown that the full-size neurotrophic factor from pigment epithelium (PEDF) induces the cell differentiation of the human promyelocyte leukemia cell line HL-60. A structural analysis of PEDF revealed in its C-terminal region a six-membered peptide fragment PEDF-(352-357) (PEDF-6) whose sequence is highly homologous to the 41-46 fragment of the active site of the human leukocyte differentiation factor HLDF (HLDF-6). The biological effect of PEDF and synthetic peptides PEDF-6 and HLDF-6 on the HL-60 cells and the early gastrula ectoderm of Xenopus laevis embryos was studied. On the basis of the structural and functional homologies of HLDF, PEDF, and their homologous peptides and the computer models of the spatial structures of the full-size PEDF and the PEDF with the C-terminal fragment split off tby the cleavage of the Leu380-Thr381 bond in the serpin loop, a hypothesis on the functional role of the serpin loop in PEDF was put forward.

Amino Acid Sequence↗
PubMed1999

A novel 45 kDa secretory protein from rat olfactory epithelium: primary structure and localisation.

cDNA clones encoding the 45 kDa protein were isolated from a rat olfactory epithelium cDNA library and their inserts were sequenced. The reconstructed protein sequence comprises 400 amino acids with a calculated molecular mass of 46,026 Da. A homology was revealed between the amino acid sequence of the 45 kDa protein and the proteins involved in the transfer of hydrophobic ligands. Using in situ hybridisation, the 45 kDa protein mRNA expression was detected in the layer of supportive cells of olfactory epithelium, apical region of trachea, surface layer of the ciliated bronchial epithelium in lung and in skin epidermis.

Amino Acid Sequence↗
PubMed1999

Localization of 28-kDa peroxiredoxin in rat epithelial tissues and its antioxidant properties.

Peroxiredoxins are a novel family of antioxidant proteins that specifically prevent enzymes from metal-catalyzed oxidation. The localization of a member of the mono-cystein subfamily of peroxiredoxins, the 28-kDa protein, in different rat tissues and its antioxidant properties were investigated. By immunoblotting, the 28-kDa peroxiredoxin was found to be most highly concentrated in olfactory epithelium and present in all tissues tested (skin, lung, trachea, kidney, womb, and brain). Immunostaining with rabbit polyclonal antibody raised against the 28-kDa peroxiredoxin revealed the particularly high level of the 28-kDa peroxiredoxin immunoreactivity in air-contacting areas (apical regions and mucus of the olfactory and respiratory epithelium and skin epidermis), which are continually exposed to numerous air-borne reactive oxygen species. In the apical regions of the olfactory and respiratory epithelium, the 28-kDa-peroxiredoxin immunogold labeling outlined microvilli and cilia and was mainly located in sustentacular cells and in respiratory and goblet cells, as electron-microscopic analysis revealed. In skin epidermis, the 28-kDa peroxiredoxin immunoreactivity was confined to the granular layer and specifically concentrated in sebaceous glands of hair follicle. In situ hybridization with 33P-labeled antisense RNA probe revealed the expression of the 28-kDa peroxiredoxin mRNA in tissues with a high level of the 28-kDa peroxiredoxin immunoreactivity. Immunodepletion of the 28-kDa peroxiredoxin profoundly decreased the antioxidant activity of the olfactory tissue extract.

Animals↗
PubMed1999

Study of immunosuppressive activity of a synthetic decapeptide corresponding to an ACTH-like sequence of human immunoglobulin G1.

The synthetic ACTH-like decapeptide H-Val-Lys-Lys-Pro-Gly- Ser-Ser-Val-Lys-Val-OH, corresponding to amino acid residues 11-20 of the variable part of the human IgG1 heavy chain (referred to as immunocortin) was found to have an immunosuppressive effect on cells in vitro: it inhibits blast transformation of mouse thymocytes and reduces spontaneous motility of mouse peritoneal macrophages as well as their bactericidal activity against the virulent bacterial strain Salmonella typhimurium 415. Tritium-labeled immunocortin binds with high affinity to ACTH receptors on thymocytes and macrophages (Kd 2. 1 and 2.5 nM, respectively) and activates adenylate cyclase in these cells. Thus, the interaction of immunocortin with the target cell includes the following main steps: binding to the receptor, activation of adenylate cyclase, and elevation of the intracellular content of cAMP.

Adenylyl Cyclases↗
PubMed1999

[Point amino acid substitutions in the Ca2+-binding centers of recoverin. I. Mechanism of successive filling of Ca2+-binding centers].

The molecule of photoreceptor Ca(2+)-binding protein recoverin contains four potential Ca(2+)-binding sites of the EF-hand type, but only two of them (the second and the third) can actually bind calcium ions. We studied the interaction of Ca2+ with recoverin and its mutant forms containing point amino acid substitutions at the working Ca(2+)-binding sites by measuring the intrinsic protein fluorescence and found that the substitution of Gln for Glu residues chelating Ca2+ in one (the second or the third) or simultaneously in both (the second and the third) Ca(2+)-binding sites changes the affinity of the protein to Ca2+ ions in different ways. The Gln for Glu121 substitution in the third site and the simultaneous Gln substitutions in the second (for Glu85) and in the third (for Glu121) sites result in the complete loss of the capability of recoverin for a strong binding of Ca(2+)-ions. On the other hand, the Gln for Glu85 substitution only in the second site moderately affects its affinity to the cation. Hence, we assumed that recoverin successively binds Ca(2+)-ions: the second site is filled with the cation only after the third site has been filled. The binding constants for the third and the second Ca(2+)-binding sites of recoverin determined by spectrofluorimetric titration are 3.7 x 10(6) and 3.1 x 10(5) M-1, respectively.

Amino Acid Substitution↗
PubMed1999

[L-glutamic acid--a modulator of the physiological status of myeloid series blood cells].

The effects of L-glutamic acid on the differentiation of HL-60 and K-562 cell lines and on the expression level of mRNAs encoding IL-1 beta, IL-6, and TNF alpha were studied. It was shown that TNF alpha actively induces the differentiation of these cell lines, whereas L-glutamic acid enhances its effect. Our results indicated that L-glutamic acid modulates the physiological state of the myeloid cell line in blood, in particular, by affecting both the reception and expression of cytokines functionally important for these cells.

Bone Marrow Cells↗