Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P30536 (PBS)
 9,886 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A high-avidity human antibody (Ab) to double-stranded DNA (dsDNA) could be virtually completely dissociated from Crithidia luciliae kinetoplast dsDNA at pH 12 and a low-avidity Ab at pH 11. At low pH (pH 2), no dissociation occurs of either Ab. Low-avidity Ab could be dissociated at neutral pH with 1 M NaCl, but even with 5 M NaCl high avidity Ab could not be dissociated. Prolonged exposure to pH 12 did not affect DNA-binding by Ab after dialysis against PBS. A decrease in surface tension of the liquid medium in no case prompted dissociation. Contact angle measurements on DNA showed it to be very hydrophilic. It is concluded that concomitant with the strong negative charge of the antigen and the positive charge of the Ab, both antigenic determinant and antibody-active site are exceptionally hydrophilic, which causes their van der Waals attraction in aqueous media to be negligibly small. This particular antigen-antibody bond thus is mainly electrostatic and can be completely dissociated by abolishing the positive charge of the antibody-active site through a drastic increase in pH.
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PMID:Dissociation of DNA/anti-DNA complexes at high pH. 700 Jun 78

Oxyrrhis marina (Dujardin) is a predatory marine dinoflagellate that feeds phagocytically on live phytoplanktonic "prey" cells from the surrounding environment. A rapid method was developed to separate the cell cycle characteristics of these predators from their prey cells in order to study the cell cycle dynamics of this organism. Nuclei from Oxyrrhis were isolated in low salt buffer (PBS) using detergent and mechanical agitation and the DNA stained with Hoechst 33258 in a one step procedure. The method permitted the isolation of nuclei from the Oxyrrhis cells with > 95% efficiency. Discrimination between prey cell nuclei and those of Oxyrrhis was achieved during flow cytometric analysis which yielded routinely G1 CVs of 3-6% for exponentially growing cell populations and 2-3% for stationary phase cells. The method was used to demonstrate the changes in cell cycle dynamics during the exponential and stationary phases of growth. Results indicated that in contrast to most mammalian and phytoplankton cell types Oxyrrhis spent the major portion (ca. 50%) of its cell cycle in G2 + M when actively dividing. Analysis of stationary phase populations also suggests that specific cell cycle control (or restriction) points were present in both G1 and G2 in this species.
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PMID:Rapid method for cell cycle analysis in a predatory marine dinoflagellate. 750 24

Formalin, an excellent preservative of cellular morphology, is a commonly used fixative for tissue specimens in hospital pathology laboratories. This preserved material is a potential source of tissue for diagnostic and retrospective research studies on DNA using flow cytometry. Unfortunately, formalin interferes with the binding of propidium iodide (PI) and other fluorescent dyes to DNA, thus altering the measurement of DNA content by flow cytometry or image analysis. This interference has been attributed to the cross-linking of histones by formalin. Since formalin alters the measurement of DNA content in formalin-fixed and formalin-fixed, paraffin-embedded tissues, this study was designed to explore the use of various physicochemical methods to reverse the effect of the formalin on the binding of PI to DNA. This study demonstrates that resuspending formalin-fixed cells in PBS and heating them at 75 degrees C for at least 1 h prior to staining with PI restores the staining of the DNA to approximately the same fluorescence intensity as that of fresh tissue.
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PMID:Reversing the effect of formalin on the binding of propidium iodide to DNA. 752 17

The Bacillus subtilis bacteriophages PBS-1 and PBS-2 protect their uracil-containing DNA by expressing an inhibitor protein (UGI) which inactivates the host uracil-DNA glycosylase (UDGase) base-excision repair enzyme. Also, PBS1/2 UGI efficiently inactivates UDGases from other biological sources, including the enzyme from herpes simplex virus type-1 (HSV-1). The crystal structure of the HSV-1 UDGase-PBS1 UGI complex at 2.7 angstrum reveals an alpha-beta-alpha sandwich structure for UGI which interacts with conserved regions of UDGase involved in DNA binding, and directly mimics protein-DNA interactions observed in the UDGase-oligonucleotide complex. The inhibitor completely blocks access to the active site of UDGase, but makes no direct contact with the uracil-binding pocket itself.
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PMID:Nucleotide mimicry in the crystal structure of the uracil-DNA glycosylase-uracil glycosylase inhibitor protein complex. 755 46

Wegener's granulomatosis (WG) is a granulomatous necrotizing vasculitis associated with the presence of ANCA, predominantly directed against proteinase 3 (PR3). The titres of ANCA correlate with disease activity and titre increases may precede disease exacerbations. Previously, we have shown that it is possible to induce autoimmune disease (systemic lupus erythematosus (SLE) and anti-phospholipid syndrome) in naive mice following active immunization with human autoantibodies, namely anti-DNA and anti-cardiolipin, respectively. The mice developed first anti-autoantibodies and, after about 4 months anti-anti-autoantibodies (Ab3), simulating auto-antibodies (Ab1) in their binding activities, and their presence was associated with the development of disease manifestations, characteristic of the human disease. So far, there is no good animal model for WG. In the current study we have immunized mice with human ANCA with the aim of inducing experimental WG. In two separate studies 30 mice were immunized in their footpads with autoantigen-purified IgG fraction (ANCA) from the sera of two patients with untreated WG, emulsified in Freund's complete adjuvant, followed 3 weeks later by ANCA injection in PBS. In the first experiment mice immunized with ANCA developed sterile microabscesses in the lungs after 8 months, and died after 8-15 months. In the second experiment, mice immunized with ANCA developed after 4 months mouse ANCA, with specificity both to PR3 and to myeloperoxidase, as well as anti-endothelial autoantibodies (AECA), as shown by radioimmunoprecipitation. Pathologically, the immunized mice developed proteinuria but not haematuria, and histological sections of the lungs demonstrated mononuclear perivascular infiltration, while diffuse granular deposition of immunoglobulins was noted in the kidneys. Our results point to a pathogenic role of ANCA in WG, and confirm the importance of the idiotypic network in the etiopathogenesis of autoimmune conditions.
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PMID:Immunization with anti-neutrophil cytoplasmic antibody (ANCA) induces the production of mouse ANCA and perivascular lymphocyte infiltration. 755 78

Reverse transcription of a retroviral RNA genome requires two template jumps to generate the linear double-stranded DNA required for integration. The RNase H activity of reverse transcriptase has several roles during this process. We have examined RNase H cleavages that define the maximal 3' and 5' ends of Moloney murine leukemia virus minus strand DNA prior to the second template jump. In both the endogenous reaction and on model substrates in vitro, RNase H cleaves the genomic RNA template between the second and third ribonucleotides 5' of the U5/PBS junction, but other minor cleavages between 1 and 10 nucleotides 5' of this junction are also observed. Similar experiments examining the specificity of RNase H for tRNA primer removal revealed that cleavage generally leaves a ribo A residue at the 5' end of minus strand DNA. These observations suggest that three bases are typically duplicated on the ends of the minus strands, leading to an intermediate following the second jump which contains unpaired nucleotides. Model substrates mimicking the structure of this intermediate demonstrate that reverse transcriptase has little difficulty in utilizing such a branched structure for the initiation of displacement synthesis.
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PMID:Cleavage specificities of Moloney murine leukemia virus RNase H implicated in the second strand transfer during reverse transcription. 759 16

Flow cytometry (FCM) is useful for measuring DNA content as related to cell cycle position. We have extended this technology by developing a method that measures cellular DNA content using propidium iodide after permeabilization with lysolecithin. This technique maintains cell integrity such that high quality mRNA can be isolated from a sorted population. Unfixed MDA-468 cells, a human breast cancer cell line, were temporarily permeabilized with lysolecithin. A range of lysolecithin concentrations were studied in order to optimize DNA staining but minimize alterations in cell size and integrity. Cells permeabilized with lysolecithin in PBS, were stained with propidium iodide (50 micrograms/ml) in the presence of 1% bovine serum albumin, 1 mM Na2EDTA in PBS. The optimal concentration (4 micrograms lysolecithin/ml) combined staining approximately 90% of the cells for DNA, with minimal effects on cell size. Subsequently, the cells were assayed for DNA content as a measure of cell cycle position. MDA-468 cells identified as being in G1 were sorted and collected. From these, high quality mRNA could be isolated, as judged by its ability to be in vitro translated into protein of a wide range of molecular weights. This technique should be useful for molecular studies requiring discrete cell populations based on DNA content and/or cell cycle position.
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PMID:Isolation of high quality mRNA from a discrete cell cycle population identified using a nonvital dye and fluorescence activated sorting. 768 93

Glucocorticoids are known to decrease wound healing by inhibiting the inflammatory response and collagen synthesis. In patients on steroids requiring emergency surgery, a safe agent that can be administered systemically is needed to reverse the deleterious effects of corticosteroids. TGF-beta and PDGF work topically but are not candidates for systemic administration. IGF-I and IGF-I:BP-3 are logical choices for systemic administration to improve wound healing. Both have been found in our laboratory to repair the corticosteroid-induced defect in wound healing when applied topically; the IGF-I:BP-3 complex gave significantly better results than IGF-I alone. Therefore, we asked whether these agents administered systemically could reverse the impaired wound healing caused by corticosteroids and whether the IGF-I:BP-3 complex was superior. Sprague-Dawley rats 350 g had 4 Hunt-Schilling wire mesh wound cylinders implanted s.c. on the back. Depo-Medrol (8 mg) was given s.c. at the time of surgery. Experimental rats were given daily s.c. injections of IGF-I or IGF-I:BP-3 (supplied by Celtrix Pharm, Santa Clara, CA) in PBS and 0.1% rat serum albumin, pH 6.0. The groups were: vehicle; IGF-I 125 micrograms/d; IGF-I:BP-3 complex containing 125 micrograms IGF-I/d. On post-op. day 17, the tissue in the wound cylinders was harvested and dried at 37 degrees C. Dry weight, DNA, total protein, and hydroxyproline (collagen) contents were obtained by our published procedures. Wound cylinder dry weight, DNA, total protein and hydroxyproline were increased by IGF-I 250%, 340%, 200% and 205%, respectively, over controls.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:In vivo effects of systemic insulin-like growth factor-I alone and complexed with insulin-like growth factor binding protein-3 on corticosteroid suppressed wounds. 768 31

The suitability of bromodeoxyuridine (BrdU) immunohistochemistry for the study of myosatellite cell proliferation in three subadult carp (Cyprinus carpio) stages (11, 15, 17 cm) was examined. They were injected intraperitoneally with BrdU and fixed one hour late. After fixation and dehydration, white myotomal muscle and small intestine samples were embedded in Histowax. Cross sections mounted on glass slides, were incubated with monoclonal antiBrdU antibody (1:100) after HCl denaturation. After washing twice in PBS, slides were incubated in goat antimouse IgG FITC secondary antibody (1:20). Single cell suspensions were obtained from gut samples. Cellular DNA partially denatured with 2 N HCl, were immunolabelled with monoclonal antibodies against BrdU. Bivariate distribution of BrdU/total cellular DNA content was measured by flow cytometry. Good visualization of BrdU labelled myosatellite cells (4-6%) and small intestine (8-9%) was obtained. Flow cytometric bivariate BrdU/DNA analysis gave evidence of the same proliferative rate in the gut samples. The applicability of these methods to fish tissues further extend the broad range of biological and biomedical investigation in which BrdU immunohistochemistry has been used.
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PMID:Use of 5'-bromodeoxyuridine immunohistochemistry to examine proliferative activity of fish tissues. 768 4

A xenograft system was developed for studying experimental carcinogenesis of human transitional cells in vivo. Segments of human ureters were tied to an injection port, ligated at the opposite end, implanted into gamma-irradiated nude rats and weekly irrigated through the injection port with fresh PBS solution. Such implants maintained the normal histologic appearance of human urothelium for at least 20 weeks in vivo in the nude rats. In situ hybridization with a human repetitive sequence DNA probe showed that the urothelium and the submucosal connective tissue and smooth muscle were of human origin. The urothelial lining was positive with an immunohistochemical reaction for acidic cytokeratins. This model allows for the long-term direct exposure of human urothelium to bladder carcinogens for the purpose of induction of human transitional cell tumors.
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PMID:A nude rat model for in vivo studies of human transitional cell carcinogenesis. 768 27


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