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)

Procedures are described for preparing monomeric selectively S-carboxamido-methylated and S-aminoethylated derivatives of seminal ribonuclease. The main properties of the derivatives, including their extinction coefficients, have been determined. Their catalytic activities and that of the S-carboxymethyl derivative have been tested. On double-stranded RNA as a substrate the monomeric derivatives are less active than the native dimeric enzyme, but much more active than pancreatic ribonuclease. On yeast RNA as a substrate the amino-ethyl derivative is found to be less active (80%) than the native enzyme, while the other two are over 30 percent more active. The monomers are stable in solution and when lyophilized from acetic acid solution do not associate to the same extent as pancreatic or native seminal ribonucleases.
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PMID:Monomeric selectively S-alkylated derivatives of seminal ribonuclease: preparation and properties. 56 95

Classically, one can increase the titer of an agglutinating or first antibody with an antiglobulin or second antibody. We have used an avidin-biotin system in place of antiglobulin to similarly extend the agglutination by an initial anticellular antibody. Erythrocytes were agglutinated by adding in succession, caproylamidobiotin-antibody, avidin, and extender (caproylamidobiotin-macromolecule). The macromolecules evaluated as extenders, in order of increasing potency, were fibrinogen, albumin, succinylated polylysine, and ribonuclease A. From comparative testing, we found that antiglobulin raised the titer of antibody from 2560 to 20,480, and the avidin-biotin tool raised the titer of caproylamidobiotin-antibody from 2560 to 10,240 without extender and to 81,820 with an extender of caproylamidobiotin-ribonuclease. Thus noncovalent extenders add to the capability of the avidin-biotin system to facilitate and substitute for an antibody.
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PMID:Enhancement of immune cellular agglutination by use of an avidin-biotin system. 57 47

We studied the uptake of modified forms of bovine pancreatic ribonuclease A (labeled with 125-iodine) by rat liver in vivo. On one hand, these experiments were intended to investigate a possible role of sinusoidal cells in the uptake of plasma proteins; on the other hand, the effect of well-defined modifications of the enzyme on the role of uptake might give us a key to the factors determining life time of plasma proteins. We used nephrectomized rats in most experiments to avoid uptake by the kidneys. Preparations of ribonuclease oligomers prepared by cross-linking with dimethyl-suberimidate enabled us to study a possible relation between uptake and molecular size. This modification does not lead to changes in charge of the protein. Monomer, dimer and polymer fractions were isolated by gel filtration on Sephadex G-75. Of the 11 amino groups in ribonuclease A, 9, 8 and 6 remained unaltered in the monomer, dimer and polymer fraction, respectively. The maintenance of biological activity, the stability of disulfide bonds, and the unchanged susceptibility to endoproteases of the cross-linked products established that gross conformational changes had not occurred. At 1 h after injection, 1% of themonomer, 7% of the dimer and 19% of the polymer were recovered per g of liver protein. Combination of autoradiography, subcellular fractionation, and the determination of labeled ribonuclease derivatives in the spleens showed that the dimer and polymer fractions were mainly present in the lysosomes of sinusoidal cells.
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PMID:Endocytosis and breakdown of proteins by sinusoidal liver cells. 61 22

Lactose has been coupled to the lysine residues of the cross-linked dimer of bovine pancreatic ribonuclease A by reductive amination with cyanoborohydride. Derivatives of ribonuclease dimer that contained up to 10 Nepsilon-1-(1-deoxylactitolyl)-lysine residues per molecule had greater than 75% of the enzymic activity of the unmodified enzyme toward yeast RNA. Upon intravenous injection of the 14C-labeled (enzymically inactivated by 14C-carboxymethylation) derivatives into rats, their uptake by the liver was a function of the number of lactose residues coupled. At 10 min, 69% of the injected derivative of ribonuclease dimer containing eight 1-deoxylactitolyl-lysine residues/molecule was found in the liver; with the non-glycosylated enzyme, the liver uptake at 10 min was only 4%, and 75% of the radioactivity was found in the kidneys.
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PMID:Effect of reductive lactosamination on the hepatic uptake of bovine pancreatic ribonuclease A dimer. 63 57

A description is given of the synthesis by fragment condensation of the peptide Gly-Glu-Ser-Arg-Glu-Ser-Ser-Ala-Asp-Lys-Phe-Lys-Arg-Gln-His-Met-Asp-Thr-Glu-Gly-Pro-Ser-Lys corresponding to the 1--23 amino acid sequence of rat pancreatic ribonuclease. This rat peptide combined with bovine S-protein yields a fully active ribonuclease S' analogue.
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PMID:Studies on polypeptides. XXVI. Synthesis of the N-terminal 1--23 peptide sequence of rat pancreatic ribonuclease; enzymatic activity of the hybrid complex with bovine S-protein. 64 56

Red kangaroo (Macropus rufus) ribonuclease was isolated from pancreatic tissue by affinity chromatography. The amino acid sequence was determined by automatic sequencing of overlapping large fragments and by analysis of shorter peptides obtained by digestion with a number of proteolytic enzymes. The polypeptide chain consists of 122 amino acid residues. Compared to other ribonucleases, the N-terminal residue and residue 114 are deleted. In other pancreatic ribonucleases position 114 is occupied by a cis proline residue in an external loop at the surface of the molecule. Other remarkable substitutions are the presence of a tyrosine residue at position 123 instead of a serine which forms a hydrogen bond with the pyrimidine ring of a nucleotide substrate, and a number of hydrophobichydrophilic interchanges in the sequence 51-55, which forms part of an alpha-helix in bovine ribonuclease and exhibits few substitutions in the placental mammals. Kangaroo ribonuclease contains no carbohydrate, although the enzyme possesses a recognition site for carbohydrate attachment in the sequence Asn-Val-Thr (62-64). The enzyme differs at about 35-40% of the positions from all other mammalian pancreatic ribonucleases sequenced to date, which is in agreement with the early divergence between the marsupials and the placental mammals. From fragmentary data a tentative sequence of red-necked wallaby (Macropus rufogriseus) pancreatic ribonuclease has been derived. Eight differences with the kangaroo sequence were found.
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PMID:The amino-acid sequence of kangaroo pancreatic ribonuclease. 65 39

Highly purified tRNAPhe from barley embryos was completely digested with pancreatic ribonuclease and T1 ribonuclease. The digestion products were separated using DEAE-cellulose chromatography. The Y base-containing fragment of the anticodon region of tRNAPhe has the following nucleotide sequence: Cpm2(2)GppsipCpApGpApCmpUpGmpApApYpAppsipCpUpGp, i.e. the same as in the anticodon region of wheat germ and pea tRNAPhe.
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PMID:Nucleotide sequence of the anticodon region of barley embryo phenylalanine transfer RNA. 66 78

Totally reduced and denatured seminal ribonuclease was regenerated using the glutathione redox system. The refolding kinetics of the enzyme were determined as a function of redox state, temperature from 14 to 43 degrees C, pH, and protein concentration. The maximal rate of regeneration occurred with 3 x 10(-3) M reduced glutathione, 6 x 10(-4) M oxidized glutathione, 24 to 30 degrees C, and pH 8.2. The products of the refolding process were characterized by Sephadex G-75, sodium dodecyl sulfate gel electrophoresis, enzymatic activity, circular dichroism, and amino acid analysis. The results indicate that the native dimeric form of the enzyme is not produced during refolding to any appreciable extent; rather, the major product is monomeric. The purified monomer exhibits twice the activity of the native enzyme toward yeast RNA. Its circular dichroism spectrum is different from the native enzyme and is quite similar to that of pancreatic ribonuclease A. Amino acid analyses showed that two glutathione molecules are bound to the monomer, suggesting that cysteine-31 and -32, which normally form the intermolecular disulfide bonds, are blocked.
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PMID:Glutathione-facilitated refolding of reduced, denatured bovine seminal ribonuclease: kinetics and characterization of products. 67 33

The dimethyl ester of bovine pancreatic ribonuclease-A (dimethyl RNAase-A), the initial product of esterification of RNAase-A in anhydrous methanolic HCl, was isolated in a homogeneous form. The two carboxy functions esterified in this derivative are those of glutamic acid-49 and aspartic acid-53. There were no changes in the u.v.-absorption spectral characteristics, the accessibility of the methionine residues, the resistance of the protein to proteolysis by trypsin and the antigenic behaviour of RNAase-A as a result of the esterification of these two carboxy groups. Dimethyl RNAase-A exhibited only 65% of the specific activity of RNAase-A, but still had the same K(m) value for both RNA and 2':3'-cyclic CMP. However, the V(max.) was decreased by about 35%. On careful hydrolysis of the methyl ester groups at pH9.5, dimethyl RNAase-A was converted back into RNAase-A. Limited proteolysis of dimethyl RNAase-A by subtilisin resulted in the formation of an active RNAase-S-type derivative, namely dimethyl RNAase-S, which was chromatographically distinct from dimethyl RNAase-A and had very nearly the same enzymic activity as dimethyl RNAase-A. Fractionation of dimethyl RNAase-S by trichloroacetic acid yielded dimethyl RNAase-S-protein and dimethyl RNAase-S-peptide, both of which were inactive by themselves but regenerated dimethyl RNAase-S when mixed together. Dimethyl RNAase-A-peptide was identical with RNAase-S-peptide. RNAase-S-protein could be generated from dimethyl RNAase-S-protein by careful hydrolysis of the methyl ester groups at pH9.5. The interaction of dimethyl RNAase-S-protein with RNAase-S-peptide appears to be about 4-fold weaker than that between the RNAase-S-protein and RNAase-S-peptide. Conceivably, the binding of the S-peptide ;tail' of dimethyl RNAase-A with the remainder of the molecule is similarly weaker than that in RNAase-A, and this brings about subtle changes in the geometrical orientation of the active-site amino acid residues of these modified methyl ester derivatives. It is suggested that these changes could be responsible for the generation of the catalytically less-efficient RNAase-A and RNAase-S molecules (dimethyl RNAase-A and dimethyl RNAase-S respectively).
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PMID:Structure and enzymic activity of ribonuclease-A esterified at glutamic acid-49 and aspartic acid-53. 70 73

The interaction between bovine pancreatic ribonuclease A (EC 3.1.4.22) and the purine nucleotides AMP, GMP, 6-chloropurine 5'-ribonucleotide and 8-bromoadenosine 5'-monophosphate was studied by u.v. difference spectroscopy. The stoicheiometry of the binding of the halogenated nucleotides to the enzyme shows a 1:1 ratio, as for the natural ones. The binding constants, Ka, for all four nucleotides at pH 5.5 were determined. They are within the same order of magnitude, though the nucleotides with a 6-amino group show a stronger interaction. The magnitude of the binding shows a reciprocal dependence on the ionic strength, which indicates an electrostatic interaction between ligand and enzyme. Finally, solvent-perturbation experiments show that all four nucleotides bind to the enzyme in a partially hydrophobic region. It is concluded that both halogenated and natural purine ribonucleotides interact in a similar manner with the enzyme molecule. The special synthesis and identification of 6-chloropurine 5'-ribonucleotide are discussed extensively. It is concluded that both halogenated and natural purine ribonucleotides interact in a similar manner with the enzyme molecule and thus the halogenated analogues are potential reagents for the affinity labelling of the purine-binding site.
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PMID:The specificity of the interaction of bovine pancreatic ribonuclease A with natural and halogenated purine nucleotides. 73 94


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