Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.31.1 (micrococcal nuclease)
 2,818 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have developed a competitive enzyme-linked immunosorbent assay for solubilized kinetochore components, using human CREST (calcinosis, Raynaud's phenomenon, esophageal dysfunction, sclerodactyly, telangiectasia) scleroderma autoimmune antibodies specific for these kinetochore elements. Using this quantitative assay, we found interphase persistent or "pre-kinetochore" components in low- and moderately high-salt (375 mM salt) extracts of micrococcal nuclease-digested rat liver and chicken erythrocyte nuclei. The release of antigen activity from nuclei under these conditions has been correlated with loss of pre-kinetochore foci as determined by immunofluorescence microscopy. Combined biochemical and competition assay analysis of chicken erythrocyte nuclear extracts indicates that pre-kinetochore components are tightly bound to chromatin of mononucleosome size. The conclusions based on competition assay data are supported by a direct binding assay, which confirms that antigens recognized by CREST sera are present on chromatin. These results raise the possibility that the kinetochore-specific chromosomal antigen(s) we have detected substitutes for "standard" mononucleosome components, such as histone H1. Furthermore, they suggest approaches to the isolation of kinetochore-specific DNA sequences from higher eucaryotes.
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PMID:Kinetochore components recognized by human autoantibodies are present on mononucleosomes. 258 Feb 29

We have partially isolated the kinetochore and associated centromeric structures from mammalian metaphase chromosomes. Human autoantibodies from scleroderma CREST (calcinosis, Raynaud's phenomenon, esophageal dysmotility, sclerodactyly, telangiectasia) patients were used as immunofluorescent probes to monitor fractionation. The procedure includes digestion of total chromosomal DNA with micrococcal nuclease, dehistonization with heparin, and dissociation of the remaining material with detergent and urea. We used a density gradient (metrizamide) to obtain an enriched fraction of stained material (kinetochore). When examined by electron microscopy, the kinetochore fraction is seen to contain numerous small immunoperoxidase-positive masses which are morphologically similar to the centromere/kinetochore region of intact metaphase chromosomes. The particulate fraction that contains kinetochore components represents less than 5% of total chromosomal proteins and contains less than 1% of total DNA. Two polypeptides of 18 and 80 kD were identified as kinetochore antigens by immunoblotting with CREST antiserum. In this paper we discuss the distribution of these kinetochore polypeptides with the associated centromeric chromatin.
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PMID:Fractionation and initial characterization of the kinetochore from mammalian metaphase chromosomes. 389 44

Antibodies directed against centromeric chromatin characteristically occur in the sera of patients with the CREST variant of scleroderma. We have studied the in situ enzymatic sensitivity and solubility of the centromeric antigen and have isolated an antigenic moiety that reacts with anticentromere antibodies. The centromeric antigen in the human epithelial cell line, HEp-2, was sensitive to DNAase I and micrococcal nuclease but not affected by RNAase A, trypsin or amylase. It was insoluble in 0.15-4 M NaCl but was extracted from the HEp-2 cells by 4 M urea/2 M NaCl. Antigenic activity in a 4 M urea/2 M NaCl extract of rabbit thymus was demonstrated by immunoabsorption. Indirect immunofluorescence of the extract separated by polyacrylamide gel electrophoresis revealed a fluorescent band with a mol. wt of 33,000. Calf thymus and trout testis histone preparations were fractionated by gel electrophoresis and transferred by blotting techniques to diazobenzyloxymethyl cellulose paper for autoradiography. Anticentromere antibodies bound to and were absorbed by trout testis histone 1. We propose that the centromeric antigen may be a variant of histone 1 that is associated with condensed chromatin.
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PMID:Anticentromere antibodies bind to trout testis histone 1 and a low molecular weight protein from rabbit thymus. 638 63