Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.27.5 (RNase)
17,967 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have reported previously [Sakakibara, et al. (1991) Chem. Pharm. Bull. 39, 146-149] that a protein purified from a partially purified pharmaceutical preparation of human chorionic gonadotropin (a urinary protein preparation from pregnant women) is a unique nonsecretory ribonuclease (RNase)-like protein on the basis of its amino terminal sequence homology. We purified the protein further from the same materials by gel filtration and reversed-phase column chromatographies with RNase activity as an index. The purified protein was designated RNase UpI-2. The catalytic activity and its sensitivity to inhibition by divalent cations suggest that the protein is related to nonsecretory RNase. The estimated molecular weight of RNase UpI-2 (38 kDa) by sodium dodecyl sulfate-polyacrylamide gel electrophoresis was significantly higher than that of urinary nonsecretory RNases (13 to 19 kDa) reported so far. After trifluoromethanesulfonic acid treatment, the molecular weight of RNase UpI-2 was reduced and approached that of nonsecretory RNase, which indicated that the protein contains a significant amount of carbohydrate (approximately 50%). RNase UpI-2 was immunoreactive with antibodies to a nonsecretory RNase, RNAase 1 [Yasuda et al. (1988) Biochim. Biophys. Acta 965, 185-194]. By immunoblot analysis of the protein freshly prepared from various urine samples, it was shown that a considerable amount of RNase UpI-2 is present in urine of pregnant women, but only a trace of RNase UpI-2, if any, was detected in urine of nonpregnant women and men. These results suggest the possibility that RNase UpI-2 may have been formed via a specific protein modification in pregnant women.
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PMID:Characterization of a unique nonsecretory ribonuclease from urine of pregnant women. 158 93

Eosinophil-derived neurotoxin (EDN) and human liver RNase were found to be indistinguishable from each other but distinct from the pancreatic ribonucleases in their nucleolytic activity on polynucleotides or small defined substrates. Antibodies to EDN and liver RNase showed identical cross-reactivities in assays of nuclease inhibition and in a radioimmunoassay. In each instance, EDN and liver RNase were easily distinguished from bovine or human pancreatic RNase. When injected intrathecally into rabbits, 5-10 micrograms of EDN or liver RNase each was neurotoxic as judged by induction of the Gordon phenomenon. Human pancreatic RNase was less neurotoxic, and up to 20-fold higher levels of bovine pancreatic RNase showed no effect. Treatment of EDN, liver RNase, and eosinophil cationic protein with iodoacetic acid at pH 5.5 resulted in inactivation of their RNase activity and also destroyed their neurotoxicity. EDN conformation was not greatly affected by iodoacetate treatment since interaction of the modified protein with antibodies was only slightly altered. We conclude that RNase activity is necessary but not sufficient to induce neurotoxic action.
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PMID:Eosinophil-derived neurotoxin and human liver ribonuclease. Identity of structure and linkage of neurotoxicity to nuclease activity. 163 26

A ribonuclease (RNase) was isolated from the urine of a 35-year-old male and purified to electrophoretic homogeneity. The enzyme was tentatively designated RNase 2. A rabbit antibody produced by injection of the purified RNase 2 was able to distinguish RNase 2 from another type of RNase coexisting in body fluids. With this antibody it was possible to detect RNase 2 isozymes in human serum and urine without difficulty using isoelectric focusing or sodium dodecyl sulfate-polyacrylamide gel electrophoresis, followed by immunoblotting. Both RNase 2 in serum and urine seemed to exist in multiple forms with regard to their molecular masses and pI values. This technique may prove to be useful in genetic and forensic studies of RNase polymorphism.
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PMID:New detection method for ribonuclease 2 (RNase 2) using immunoblotting with specific antibody. 235 60

Eosinophil-derived neurotoxin (EDN) and eosinophil cationic protein (ECP) were isolated from lysates of human eosinophil granules by gel filtration and ion exchange chromatography on heparin-Sepharose. Radioimmunoassay, using monoclonal antibodies, of fractions from the heparin-Sepharose chromatography showed one peak of EDN activity and two peaks of ECP activity (termed ECP-1 and ECP-2). EDN, ECP-1, and ECP-2 each exhibited heterogeneity in charge and molecular weight when analyzed by two-dimensional nonequilibrium pH gradient electrophoresis and NaDodSO4/PAGE. Digestion of EDN with endoglycosidase F (endo F) decreased its molecular weight and charge heterogeneity. Thus, END likely contains a single complex oligosaccharide. Endo F digestion of ECP-1 and ECP-2 decreased the molecular weight of both polypeptides, indicating that both likely contain at least one complex oligosaccharide. Amino acid sequence analyses showed that ECP-1 and ECP-2 are identical from residue 1 through residue 59 and that the sequences of EDN and ECP are highly homologous (37 of 55 residues identical). Both EDN and ECP NH2-terminal sequences showed significant homology to RNase, especially in regions of the RNase molecule involved in ligand binding. EDN, ECP-1, and ECP-2 had neurotoxic activity, causing the Gordon phenomenon at doses down to 0.15 micrograms when injected into the cisterna magna; the proteins were comparable in their activities. These results indicate that EDN and ECP are related proteins and suggest that they derived from genes associated with the RNase family.
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PMID:Biochemical and functional similarities between human eosinophil-derived neurotoxin and eosinophil cationic protein: homology with ribonuclease. 345 70

Two alkaline RNases, designated RNase 1 and RNase 2, were isolated from the culture media of silica-treated and non-treated macrophages. The yield of RNase from the medium of silica-treated macrophages was 30% of that from the non-treated control. The effects of these RNases on cultured granuloma fibroblasts and on granulation-tissue nuclei were studied. RNase 1 inhibited thymidine incorporation into fibroblasts except at low concentrations, where it was observed to be stimulatory. RNase 1 also inhibited the protein synthesis of fibroblasts. The incorporation of cytidine into RNA in cultured fibroblasts was not affected by RNase 1, but the incorporation into isolated nuclei was decreased. In pulse chase experiments RNase 1 increased the release of cytidine, but not that of thymidine, from the cells. RNase 2 had no effect on the protein or nucleic acid metabolism of the fibroblasts or on the RNA metabolism of isolated nuclei, perhaps because of impermeability. These experiments confirm that macrophage RNase activity is able to regulate the metabolism of granulation-tissue fibroblasts by increasing RNA degradation. Through this action it also regulates DNA and protein synthesis and other metabolic functions of those cells.
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PMID:Effects of purified macrophage RNase on granuloma fibroblasts with reference to silicosis. 625 52

The ribonucleases (RNases) of human blood serum, urine, cerebrospinal fluid (CSF), and leukocytes were visualized by activity staining after electrophoresis in RNA-case sodium dodecyl sulfate-polyacrylamide gels. Samples were prepared for electrophoresis by heating for 2 min at 100 degrees C in 2% sodium dodecyl sulfate (NaDodSO4) and 5% mercaptoethanol, conditions which dissociate proteins into their constituent polypeptide chains and permit estimation of molecular weight. It was found that each of the five peaks of serum alkaline RNase activity separable on phosphocellulose columns, i.e., RNases 1-5 of Akagi et al. [Akagi, K., Murai, K., Hirao, N., & Yamanaka, M. (1976) Biochim. Biophys. Acta 442, 368-378], is associated with electrophoretically distinct enzymes. The molecular weights exhibited by these enzymes in NaDodSO4 gels are 31 000 and 28 000 (major species of RNase 1), 25 000 (RNase 2), 20 000 (RNase 3), 16 000 (RNase 4), and 14 000 (RNase 5). The RNase activity of leukocytes displays a molecular weight of 17 000 and exhibits a characteristic dependence of its Rf on the temperature at which samples (in 2% NaDodSO4 without mercaptoethanol) are prepared for electrophoresis. An RNase activity like that of leukocytes, distinct from RNases 1-5, is found in serum. Urine RNase activity is less heterogeneous than that of serum, consisting mainly of species like serum RNase 1 and an enzyme similar to leukocyte RNase. Conversely, CSF RNase activity is more complex and includes enzymes resembling serum RNases 1-5 as well as additional species either not observed in serum or detected in serum as minor components following chromatography. The analytical methods described herein are particularly useful for assessment of heterogeneity of RNase preparations and for direct comparison of the RNases of crude and purified samples.
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PMID:Ribonucleases of human serum, urine, cerebrospinal fluid, and leukocytes. Activity staining following electrophoresis in sodium dodecyl sulfate-polyacrylamide gels. 723 97

Four major urine ribonuclease (RNase) activities, designated bands A-D, were identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and activity staining. Bands A, B, and C have alkaline pH optima and display molecular weights of 31 000, 23 000, and 20 000, respectively, upon sodium dodecyl sulfate (NaDodSO4) gel electrophoresis and weights of 44 000, 28 000, 22 000 upon gel filtration. Band D, with a pH optimum slightly below neutrality, has a molecular weight of 16 000 or 15 000, respectively, determined by the above methods. Band A, the most abundant activity in urine, is heterogeneous and resembles serum RNase 1 on electrophoresis and on phosphocellulose and Sephadex chromatography. Band B is similar to a minor, unnamed component of serum RNase activity while band C resembles serum RNase 3. Band D is similar to the leukocyte RNase-like activity of serum [Blank, A., & Dekker, C.A. (1981) Biochemistry (preceding paper in this issue)]. Band A is present in urine at a concentration high than that of RNase 1 in serum. In contrast, urine counterparts of serum RNases 2, 4, and 5 are not apparent upon either phosphocellulose chromatography [see also Yamanaka, M., Akagi, K., Murai, K., Hirao, N., Fujimi, S., & Omae, T. (1977) Clin. Chim. Acta 78, 191-201] or NaDodSO4 get electrophoresis; a urine counterpart of serum RNase 3 can be detected only by the more sensitive electrophoretic method. These results indicate that RNase 2-5 are processed differently by the kidney than RNase 1. After reconciliation of reported differences in their pH optima and molecular weights, five apparently diverse RNase preparations described in the literature can be related to band A activity and three preparations to band D. However, we are unable to confirm a previous report of a human urine enzyme indistinguishable from bovine pancreatic RNase A.
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PMID:Multiple ribonucleases of human urine. 723 98

A ribonuclease (RNase) that cleaves specifically on the 3' side of uridine [Shapiro, R., Fett, J. W., Strydom, D. J. & Vallee, B. L. (1986a) Biochemistry 25, 7255-7264] was purified from human plasma and its amino acid sequence was determined. This protein is a 119-residue single-chain polypeptide cross-linked by four disulfide bonds and has an amino-terminal pyroglutaminyl residue. No post-translational modifications were observed during extensive sequence studies on peptide fragments, except for the amino-terminal pyroglutamic acid and a possible deamidation of Asn66. The protein is homologous to the pancreatic ribonucleases and angiogenin, but differs substantially from both of these proteins; the protein sequence has 43% identity with human pancreatic ribonuclease and 39% identity with human angiogenin, as compared to 35% identity between human angiogenin and pancreatic ribonuclease. It is referred to as RNase 4, based on the nomenclature currently used for the genes of pancreatic RNase (RNase 1) and the eosinophil-derived RNases (RNase 2 and RNase 3). Virtually all of the RNase active-site components, including the catalytic residues His12, His119 and Lys41, are preserved. However, some invariant residues of RNase 1 are replaced, e.g. Lys7 by arginine, Asp14 by histidine, and Pro42 by arginine. RNase 4 contains a unique two-residue deletion at the position corresponding to amino acids 77 and 78 of pancreatic RNase, and its carboxyterminal sequence is truncated at position 122. The deletion in angiogenin at position 21 is also found in RNase 4. RNase 4 is very similar to two RNases isolated from bovine and porcine liver, and together they form a new family in the RNase superfamily. The degree of inter-species similarity (90%) is much greater than within the pancreatic RNase and angiogenin families, which suggests that this ribonuclease could possess a physiologically important function other than general RNA catabolism.
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PMID:The amino acid sequence of human ribonuclease 4, a highly conserved ribonuclease that cleaves specifically on the 3' side of uridine. 822 79

Eosinophil-derived neurotoxin (EDN) is a ribonuclease with neurotoxic and helminthotoxic properties. It is present in the crystalloid granules of human eosinophils. We report the expression and characterization of a functionally active recombinant human EDN using the pMAL-cRI expression system. A cDNA for mature EDN was obtained by PCR and inserted in pMAL-cRI downstream of the malE gene encoding maltose binding protein. Induction of the ptac promoter of the plasmid in Escherichia coli strain BL21(DE3) resulted in high level expression of soluble MAL-EDN fusion protein. Cleavage of affinity purified fusion protein with Factor Xa protease released recombinant EDN which comigrated with native EDN on SDS-polyacrylamide gels and cross-reacted with a polyclonal anti-EDN antiserum on Western blots. IN contrast to previous attempts at EDN expression, denatured and refolded EDN had ribonuclease activity and was prepared in microgram amounts. The availability of recombinant human EDN should facilitate studies of its structure and biological functions.
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PMID:Expression in Escherichia coli and purification of human eosinophil-derived neurotoxin with ribonuclease activity. 853 63

C2-alpha-Mannosyltryptophan was discovered in RNase 2 from human urine, representing a novel way of attaching carbohydrate to a protein. Here, we have addressed two questions related to the biosynthesis of this modification: (i) is C-mannosylation part of the normal intracellular biosynthetic route, and (ii) how general is it, i.e. which organisms perform this kind of glycosylation? To answer the first question, RNase 2, which is identical to the eosinophil-derived neurotoxin, was isolated from intracellular stores of cultured human HL-60 cells. The enzyme was C-mannosylated at Trp-7, showing that the modification occurs intracellularly, before secretion of the protein. The second question was investigated by immunological and chemical analysis of RNase 2 purified from the supernatant of transiently transformed cells from different organisms. This revealed that C-mannosylation occurs in cells from man, green monkey, pig, mouse, and hamster. The observation that pig kidney cells contain the machinery for C-mannosylation of Trp-7 of human RNase 2 but that the homologous RNase from porcine kidney is not a substrate, since it does not contain a tryptophan at position 7, strongly suggests that C-mannosylated proteins other than RNase 2 exist. Recombinant RNase 2 isolated from insect cells, plant protoplasts, and Escherichia coli was not C-mannosylated. These results not only form the basis for further studies on the biochemical aspects of C-mannosylation but also have implications for the choice of cells for production of recombinant glycoproteins.
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PMID:C-Mannosylation of human RNase 2 is an intracellular process performed by a variety of cultured cells. 933 52


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