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Biomedical subjects

R J DeLange

Publications and source records attributed to R J DeLange.

At least 19 recordsLinked to original sources
PubMed1990

Identification and partial purification of a chromatin bound calmodulin activated histone 3 kinase from calf thymus.

A calcium-calmodulin (Ca2(+)-CaM) stimulated histone H3 phosphorylating activity was identified as a component of a nuclear protein complex purified from a 150 mM NaCl extract of calf thymus chromatin. This activity bound to a CaM-Sepharose affinity column in a Ca2+ dependent manner and was eluted off the column in the presence of EGTA. Equilibrium centrifugation of the EGTA eluate on a sucrose density gradient revealed that the activity is a component of a larger complex identified at 25% sucrose. This complex consisted of two major proteins, having Mr of 65 and 75 kDa. Using [125I] CaM and the gel overlay technique it was shown that the 75 kDa protein is the major CaM binding protein in the complex.

Animals↗
PubMed1990

Bile acids: antioxidants or enhancers of peroxidation depending on lipid concentration.

Using three different assay systems, we have discovered a heretofore unrecognized antioxidant property of bile acids at physiological concentrations. Bile acids inhibit peroxidation of the polyunsaturated lipid, linoleic acid, and of the highly fluorescent protein phycoerythrin. In part, the antioxidant activity results from scavenging of peroxyl radicals by direct oxidation of the bile acids. The most abundant products of the reaction of cholate and chenodeoxycholate with peroxyl radicals were studied in detail and shown to be the keto derivatives formed by oxidation of the 7 alpha-hydroxyl groups. Paradoxically, at linoleate concentrations higher than 1-2 mM, glycocholate up to approximately 10-14 mM enhances lipid peroxidation and inhibits only at higher concentrations. These findings may prove important in understanding the etiology of certain disease states of the biliary tract and intestine where lipid peroxidation may be involved and in providing a rationale for the positive epidemiological correlation between high lipid intake and higher fecal bile acid output and colon cancer.

Antioxidants↗
PubMed1989

Phycoerythrin fluorescence-based assay for peroxy radicals: a screen for biologically relevant protective agents.

Under the conditions of this assay, antioxidants that react rapidly with peroxy free radicals (e.g., ascorbate, vitamin E analogs, urate), protect phycoerythrin completely from damage by such radicals generated by thermal decomposition of 2,2'-azobis(2-amidinopropane); other compounds provide partial concentration-dependent protection. Change in phycoerythrin fluorescence emission with time provides a measure of the rate of free radical damage. The assay exploits the unusual reactivity of phycoerythrin toward these peroxy radicals. On a molar basis, phycoerythrin reacts with these radicals over 100-fold slower than do ascorbate or vitamin E analogs, but over 60-fold faster than other proteins. Applications of this assay to the estimation of the peroxy radical scavenging capacity of human plasma are described, and to the comparison of the scavenging properties of several proteins and of DNA, of vitamins and their derivatives, of catecholamine neurotransmitters, and of a variety of other low molecular weight biological compounds.

Antioxidants↗
PubMed1988

Monoclonal antibody toward lysosomal cathepsin B cross-reacts preferentially with distinct histone classes.

1. A set of monoclonal antibodies (Mab) was prepared against cathepsin B (CB) from rat preputial-gland, an organ characterized by rapidly-renewing cell populations, which is a uniquely enriched source of lysosomal enzymes, including CB. Minute amounts of CB are known to be transferred abruptly to the nuclear compartment in a variety of activated cells. 2. Since, on the basis of its stringent substrate requirements, CB was expected to function at limited protein loci in chromatin, Mab Line II-B4 was used to probe Western blots of chromatin fractions and selected proteins. 3. The Mab, which was not directed against the active site of CB, cross-reacted preferentially with histones 3 and 4 (H3 and H4) in acid-soluble fractions of chromatin from rat preputial-gland. Line II-B4 also recognized H3 and H4 selectively in calf thymus histones and among histones purified from a wide range of sources from yeast to man. HMG 1 was minimally immunoreactive among preputial gland constituents and carbonic anhydrase (CA) was also sensitive to the Mab. 4. The common determinants were not shared by any of the H1 series, nor by H2A, H2B, protein A24 or a wide range of natural and synthetic products. 5. Origin of the antigenicity was traced by chemical modifications of H3, H4 and CA to the critical contribution of arginine and hydrophobic amino acid residues in its immediate environment, indicating that Line II-B4 may be directed against an epitope comprising the specific binding-site of CB and its selective substrate(s). 6. These data suggest that certain highly conserved cellular constituents may be uniquely vulnerable to limited proteolysis in preproliferative cells responding to mitotic signals.

Animals↗
PubMed1987

Determination of estrogen and androgen receptors in Trichomonas vaginalis and the effects of antihormones.

Trichomonas vaginalis, a common genital pathogen, was found to possess both specific estrogen and specific androgen receptors. These 4.3 S macromolecules were proteinaceous in nature. Both metronidazole-resistant and metronidazole-sensitive strains possessed both types of sex hormone-binding proteins. The estrogen receptor binding was competitively inhibited by the antiestrogen tamoxifen citrate, and the androgen binding was competitively inhibited by the antiandrogen cyoctol. The presence of these specific receptors may allow the use of hormonal and antihormonal manipulation in the treatment of infections caused by these organisms.

Androgen Antagonists↗
PubMed1985

Contact site of histones 2A and 2B in chromatin and in solution.

Irradiation of isolated nuclei or of a complex of histones 2A (H2A) and 2B (H2B) with ultraviolet light produces a covalent cross-link between H2A and H2B. Sequence analysis of the peptides isolated from the H2A-H2B dimer formed in solution and in nuclei demonstrated that both dimers are produced through the covalent linkage of Tyr-40 of H2B and Pro-26 of H2A. Tyrosyl residues proximal to Tyr-40 did not produce a cross-link with H2A, thereby indicating that strict conformational parameters are required for production of the H2A-H2B cross-link. We conclude that the precise juxtaposition of Tyr-40 of H2B and Pro-26 of H2A in this region of the H2A/H2B contact site is not altered upon interaction of these histones with H3 and H4 (tetramer), DNA, or other chromosomal components during nucleosome assembly.

Amino Acid Sequence↗
PubMed1985

Accessibility of tyrosyl residues altered by formation of the histone 2A/2B complex.

The availability of tyrosyl residues to surface iodination was analyzed for histone 2A (H2A), histone 2B (H2B), and the H2A/H2B complex. When H2A is free in solution (200 mM NaCl, pH 7.4) tyrosine-39 and one or both tyrosines-50 and -57 were readily iodinated. Tyrosines-83 and -121 of H2B were iodinated, both when the histone was free in solution and when it was associated with H2A, while tyrosines-37, -40, and -42 of H2B were not iodinated under either condition. When H2A and H2B were associated or covalently cross-linked, all tyrosyl residues of H2A were unavailable for iodination. We also found that the iodination of nondenatured H2A and H2B did not inhibit formation of the H2A/H2B complex. These results indicate that the amino-terminal regions of the hydrophobic portions of H2A and H2B undergo significant conformational changes upon formation of the H2A/H2B complex. These conformational shifts occur in the same region of the H2A/H2B complex that contains a contact site between H2A and H2B in the nucleosome, thus indicating an involvement of this region in chromatin assembly.

Animals↗
PubMed1985

Primary structures of six antimicrobial peptides of rabbit peritoneal neutrophils.

Six microbicidal peptides, NP-1, NP-2, NP-3a, NP-3b, NP-4, and NP-5, from rabbit peritoneal neutrophils were characterized. As a family, the peptides were 32-34 residues in length, were cystine- and arginine-rich, and each contained three intramolecular disulfide bonds. Within their sequences, the six peptides shared 11-residue positions, which included the six half-cystines contained in each peptide. NP-1 and NP-2 differed by a single residue and were identical in their respective sequences to MCP-1 and MCP-2, the peptide analogs from rabbit alveolar macrophages. NP-4 and NP-5 were homologous in 27 of their residues, but NP-3a and NP-3b shared little more than the 11-residue backbone common to all members of this peptide family.

Amino Acids↗
PubMed1985

Antiandrogen and antineoplastic effects of a novel drug, CPC10997.

CPC10997 was found to be effective in vitro as an antiandrogen without effects on either the estrogen or the progesterone receptors in carcinomas of the breast, ovary and prostate as well as in malignant melanomas. Using the clonogenic assay of Salmon et al. [Cancer Treat. Rep. 65: 1-12, 1981], CPC10997 was found to be more effective against carcinomas of the breast, the kidney, the ovary and the prostate than conventional antineoplastic agents in the majority of tumors tested. In view of the very favorable toxicology profiles and in vitro efficacy, further trials using CPC10997 as an antineoplastic agent are indicated.

Androgen Antagonists↗
PubMed1984

Purification of bovine bone morphogenetic protein by hydroxyapatite chromatography.

Bovine bone morphogenetic protein (bBMP) induces differentiation of mesenchymal-type cells into cartilage and bone. bBMP has an apparent Mr of 18,500 +/- 500 and represents less than 0.001% of the wet weight of bone tissue. A Mr 34,000 protein resembling osteonectin is separated by extraction with Triton X-100. A Mr 24,000 protein and about half of a Mr 22,000 protein are disassociated from bBMP by precipitation in 1.5 M guanidine hydrochloride. Aggregates of bBMP and a Mr 14,000 protein are insoluble in aqueous media; the bBMP becomes soluble when the Mr 14,000 protein is disassociated in 6 M urea and removed from the solution by ultrafiltration. Three separate molecular species with apparent Mrs 18,500, 17,500, and 17,000 are eluted at 0.10, 0.15, and 0.20 M phosphate ion concentrations, respectively, from a hydroxy-apatite column. The Mr 18,500 protein has the amino acid composition of acidic polylpeptide and includes four half-cystine residues; the pI is 4.9-5.1. The Mr 22,000 component is a chromoprotein resembling ferritin. The NH2-terminal amino acid sequence of the Mr 17,500 protein simulates histone H2B. The Mr 17,000 protein may possess calmodulin activity. Aggregates of the Mr 18,500 and other proteins induce formation of large deposits of bone; the Mr 18,500 protein alone is rapidly absorbed and induces formation of small deposits. None of the other proteins induces bone formation.

Amino Acid Sequence↗
PubMed1983

Histone-like protein in the Archaebacterium Sulfolobus acidocaldarius.

The Archaebacterium Thermoplasma acidophilum contains a basic chromosomal protein remarkably similar to the histones of eukaryotes. Therefore, it was of interest to examine a different Archaebacterium for similar proteins. We chose to examine Sulfolobus acidocaldarius because it is thermophilic, like T. acidophilum, but nevertheless the two organisms are not particularly closely related. Two major chromosomal proteins were found in S. acidocaldarius. The smaller of these was soluble in 0.2 M H2SO4 and had a molecular weight of 14500. The larger was acid-insoluble and had a molecular weight of about 36000. Together, the proteins protected about 5% of the DNA against nuclease digestion and stabilized about 50% against thermal denaturation. Overall, the properties of these proteins were intermediate between those of the Escherichia coli protein HU and T. acidophilum protein HTa.

Amino Acids↗
PubMed1983

Bone cell differentiation and growth factors.

Bone morphogenetic protein and bone-derived growth factors are biochemical tools for research on induced cell differentiation and local mechanisms controlling cell proliferation. Bone morphogenetic protein irreversibly induces differentiation of perivascular mesenchymal-type cells into osteoprogenitor cells. Bone-derived growth factors are secreted by and for osteoprogenitor cells and stimulate DNA synthesis. Bone generation and regeneration are attributable to the co-efficiency of bone morphogenetic protein and bone-derived growth factors.

Animals↗
PubMed1979

The amino acid sequence of fragment A, an enzymically active fragment of diphtheria toxin. I. The tryptic peptides from the maleylated protein.

Six tryptic peptides ranging in size from 3 to 126 residues were isolated from maleylated Fragment A of diphtheria toxin after tryptic hydrolysis. These peptides accounted for all 193 residues found by amino acid analysis. After demaleylation, the six peptides were purified by chromatography on Sephadex G-50, coupled with paper chromatography and electrophoresis, and were analyzed by various methods. The compositions and properties of the peptides are reported. Almost 70% of the residues were positioned within these peptides.

Adenosine Diphosphate Sugars↗
PubMed1979

The amino acid sequence of fragment A, an enzymically active fragment of diphtheria toxin. II. The cyanogen bromide peptides.

Cyanogen bromide cleavage of Fragment A from diphtheria toxin at the four methionines present in each molecule resulted in five major peptides which were isolated and studied by sequence methods. These five peptides of 4, 11, 14, 63, and 101 residues account for all 193 residues in Fragment A and provide overlaps for the tryptic peptides from the maleylated protein. Two additional peptides were isolated and shown to be shorter forms (8 and 10 residues) of the COOH-terminal cyanogen bromide peptide (11 residues).

Adenosine Diphosphate Sugars↗
PubMed1979

The amino acid sequence of fragment A, an enzymically active fragment of diphtheria toxin. III. The chymotryptic peptides, the peptides derived by cleavage at tryptophan residues, and the complete sequence of the protein.

Fragment A (21,145 daltons in its longest known form) may be derived from diphtheria toxin (60,000 daltons) by mild tryptic digestion and reduction. Purified Fragment A consists of a mixture of 3 molecules of 190, 192, and 193 residues; the first 190 residues are in common and correspond to the NH2-terminal region the toxin. All three species of Fragment A are active in catalyzing ADP ribosylation of elongation factor 2, an essential component of protein synthesis. This reaction inactivates the factor and is responsible for the toxin's action in inhibiting protein synthesis in animal cells. It is believed that Fragment A or similar enzymically active fragments released into the cytosol of toxin-treated cells mediate this inhibition. The complete amino acid sequence of Fragment A has been determined from 32 chymotryptic peptides, three peptides derived by chemical cleavage of Fragment A at its 2 tryptophan residues, five cyanogen bromide peptides, and six tryptic peptides from the maleylated protein.

Adenosine Diphosphate Sugars↗
PubMed1979

Identification of interacting amino acids at the histone 2A--2B binding site.

Histones 2A and 2B of calf thymus were cross-linked within intact nuclei by UV irradiation. This procedure induces the formation of covalent cross-links between noncovalently interacting residues in the histones of native chromatin. Tryptic peptide and partial sequence analysis of the cross-linked product has shown that the covalent linkage is between tyrosine-37, -40, or -42 (we have not yet determined which) of H2B and proline-26 of H2A. We conclude that these residues constitute part of the hydrophobic H2A--H2B binding domain within the nucleosomes of native chromatin.

Amino Acid Sequence↗