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:3.1.26.9 (ribonuclease)
6,589 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

One of the four titrating histidine ring C-2 proton resonances of bovine pancreatic ribonuclease has been assigned to histidine residue 12. This was accomplished by a direct comparison of the rate of tritium incorporation into position C-2 of histidine 12 of S-peptide (residues 1 to 20) derived from ribonuclease S, with the rates of deuterium exchange of the four histidine C-2 proton resonances of ribonuclease S under the same experimental conditions. The same assignment was obtained by a comparison of the NMR titration curves of ribonuclease S, the noncovalent complex of S-peptide and S-protein (residues 21 to 124) with the results for the recombined complex in which position C-2 of histidine 12 was fully deuterated. The second active site histidine resonance was assigned to histidine residue 119 by consideration of the NMR titration results fro carboxymethylated histidines and 1-carboxymethylhistidine 119 ribonuclease. This assignment is a reversal of that originally reported, and has important implications for the interpretation of NMR titration data of ribonuclease.
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PMID:Nuclear magnetic resonance titration curves of histidine ring protons. A direct assignment of the resonances of the active site histidine residues of ribonuclease. 0 54

The histidine C-2 proton NMR titration curves of ribonuclease S-peptide (residues 1 to 20) and S-protein (residues 21 to 124) are reported. Although S-protein contains 3 histidine residues, four discrete resonances are observed to titrate. One of these arises from the equivalent histidine residues of unfolded S-protein. The variation in area of the four resonances indicate that there is a reversible pH-dependent equilibrium between the folded and unfolded forms of S-protein, with some unfolded material being present at most pH values. Two of the resonances of the folded S-protein can be assigned to 2 of the histidine residues, 48 and 105, from the close similarity of their titration curves to those in ribonuclease. These similarities indicate a homology of portions of the folded conformation of S-protein to that of ribonuclease in solution. These results indicate that the complete amino acid sequence is not required to produce a folded conformation similar to the native globular protein, and they appear to eliminate the possibility that proteins fold from their NH2 terminus during protein synthesis. The low pH inflection present in the titration curve assigned to histidine residue 48 in ribonuclease is absent from this curve in S-protein. This is consistent with our previous conclusion that this inflection arises from the interaction of histidine 48 with aspartic acid residue 14, which is also absent in S-protein. The third titrating resonance of native S-protein is assigned to the remaining histidine residue at position 119. The properties of this resonance are not identical with either of the titration curves of the active site histidine residues 12 and 119 of ribonuclease. The resonance assigned to histidine 119 is the only one significantly affected on the addition of sodium phosphate to S-protein, indicating that some degree of phosphate binding occurs. In both the absence and presence of phosphate this curve also lacks the low pH inflection observed in the histidine 119 NMR titration curve in ribonuclease. This difference presumably arise from a conformational between ribonuclease and the folded S-protein involving a carboxyl group.
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PMID:Nuclear magnetic resonance titration curves of histidine ring protons. Ribonuclease S-peptide and S-proteins. 0 55

Four antigenic regions of native bovine pancreatic ribonuclease have been located by using antibodies that react specifically with segments 1--13, 31--79, and 80--124. These antibodies were purified by affinity chromatography on columns to which these peptide segments were bound. Analysis of precipitin curves indicates that there are at least three antigenic determinants to which antibody molecules can bind simultaneously in the presence of excess antibodies. Analysis of binding data, however, for each purified specific antibody preparation, carried out by the method of Berzofsky et al. [Berzofsky, J. A., Curd, J. G., & Schechter, A. N. (1976) Biochemistry, 15, 2113], leads to an estimate of four for the number of antigenic determinants in ribonuclease; this estimate had also been made earlier by Stelos et al. [Stelos, P., Fothergill, J. E., & Singer, S. J. (1960) J. Am. Chem. Soc. 82, 6034]. We find that one determinant is associated with each of segments 1--13 and 80--124 and two with segment 31--79. No antigenic activity could be detected for segment 14--29 either in native ribonuclease or in the free fragment. These conclusions are based on (1) the use of specific peptides to isolate purified antibodies by affinity chromatography, (2) immunoprecipitation of an antigenic peptide from the peptic digest of ribonuclease, (3) competitive inhibition studies with various peptide and protein fragments [cyanogen bromide fragments 1--13, 31--79, and 80--124, the tryptic peptides 40--61 and 105--224, S-peptide, S-protein, and des(121--124)-RNase], and (4) comparison and evaluation of the published effects on antigenicity of chemical and enzymatic modifications and changes in sequence among homologous ribonucleases. These approaches provide evidence that the four antigenic determinants are localized around the alpha-helical portion of segment 1--10, somewhere in segment 40--61, at the beta bend in segment 63--75, and either at the beta bend or beta sheet in segment 87--104 of native ribonuclease.
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PMID:Location of the antigenic determinants of bovine pancreatic ribonuclease. 9 May 20

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

Investigation of the known protein structures has led to the generalization that the native folding permits each sidechain to select those nearest-neighbors which maximize stabilization from van der Waals interactions. With regard to secondary structure: 1. Helical and beta regions exhibit characteristic patterns of short-range contacts (residue numbers k and k + t with [t] less than or equal to 4) due to the geometries of these secondary structures. However, these are not strictly obligatory, and preferred short-range contacts which would result in unfavorable van der Waals interactions are replaced by favorable long-range contacts. 2. The generalization mentioned at the outset holds for individual proteins, both for short-range and long-range contacts, and without regard for the type or amount of secondary structure present. 3. These observations imply that van der Waals interactions arising from short-range contacts partially determine secondary structure, and this is demonstrated by tests based upon assignment of regions of secondary structure in the known proteins. The principle of optimizing van der Waals stabilization from long-range contacts is applied to predict the structure of the complex formed by the S-peptide and S-protein of ribonuclease-S. The formation of favorable pairs is found to be more important than the total number of intermolecular contacts, and 40 to 50% of this stabilization is contributed by two residues of the S-peptide, Phe-8 and Met-13.
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PMID:Local interactions as a structure determinant for protein molecules: III. 76 Aug 7

Syntheses are described of two S-peptide analogues where the arginyl residue in position 10 has been replaced by ornithine and the phenylalanine in position 8 has been substituted by the unnatural amino acids cyclohexylalanine or p-fluorophenylalanine. In order to regenerate the arginyl residue, which is present in position 10 in the natural sequence, the S-peptide analogues beloning to the [Orn10]-series are transformed into the corresponding guanidinated derivatives by treatment with O-methylisourea. 1epsilon, 7epsilon, 10delta triguan-[Cha8, Orn10]-, 1epsilon, 7epsilon, 10delta-triguan-[pF-Phe8, Orn10]- and 1epsilon, 7epsilon, 10delta-triguan-[Tyr8, Orn10]-S-peptides were prepared. The ability to bind to and activate the S-protein of the synthetic S-peptide analogues, before and after guanidination, was tested by exploring their capacity to generate ribonuclease activity using RNA and C greater than p as substrates. The affinity of the different peptides for the S-protein in the absence of substrate was evaluated by difference spectroscopy.
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PMID:Kinetic and conformational studies on some partially synthetic ribonuclease S' analogues modified in position 8. 88 Dec 90

Limited proteolysis of RNAase-Aa(1) (monodeamidated ribonuclease-A) by subtilisin results in the formation of an active RNAase-S type of derivative, namely RNAase-Aa(1)S. RNAase-Aa(1)S was chromatographically distinct from RNAase-S, but exhibited very nearly the same enzymic activity, antigenic conformation and susceptibility to trypsin as did RNAase-S. Fractionation of RNAase-Aa(1)S by trichloroacetic acid yielded RNAase-Aa(1)S-protein and RNAase-Aa(1)S-peptide, both of which are inactive by themselves, but regenerate active RNAase-Aa(1)S' when mixed together. RNAase-Aa(1)S-peptide was identical with RNAase-S-peptide, whereas the protein part was distinct from that of RNAase-S-protein. Titration of RNAase-Aa(1)S-protein with S-peptide exhibited slight but noticeably weaker binding of the peptide to the deamidated S-protein as compared with that of native protein. Unlike the subtilisin digestion of RNAase-A, which gives nearly 100% conversion into RNAase-S, the digestion of RNAase-Aa(1) gives only a 50% conversion. The resistance of RNAase-Aa(1) to further subtilisin modification after 50% conversion is apparently due to the interaction of RNAase-Aa(1) with its subtilisin-modified product. RNAase-S was also found to undergo activity and structural changes in acidic solutions, similar to those of RNAase-A. The initial reaction product (RNAase-Sa(1)) isolated by chromatography was not homogeneous. Unlike the acid treatment of RNAase-A, which affected only the S-protein part, the acid treatment of RNAase-S affected both the S-protein and the S-peptide region of the molecule.
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PMID:Subtilisin modification of monodeamidated ribonuclease-A. 92 53

The relationship of structure to function in the recognition of ribonuclease S-peptide by S-protein was studied by several methods. Liquid phase peptide synthesis was employed to generate analogs of S-peptide in which from 1 to 8 residues were deleted from the NH2-terminal end of the S-peptide. Additional derivatives were made by substitutions in the NH2-terminal three amino acids or by modifying the S-peptide analogs by trifluoroacetylation. The analogs were generated in the following way. S-Peptide was cleaved with chymotrypsin. The fragment obtained, RNase(9-20), was purified and lengthened step by step using liquid phase peptide synthesis. A second set of analogs were prepared by cleavage of CF3CO-S-peptide with elastase and the resulting CF3CO-RNase(7-20), similarly lengthened. The various analogs of S-peptide were tested in their capacity to combine with S-protein and regenerate biological activity as measured by Vmax and Kb. This work shows a positive contribution of every one of the first 8 NH2-terminal residues of S-peptide to the molecular recognition of S-protein in the presence of RNA substrate. Substitution of the first 3 residues by alanine or blocking of the free amino groups decreases recognition, indicating that the original primary structure is the most favorable one.
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PMID:Ribonuclease S-peptide. A model for molecular recognition. 125 70

A defective S-allele, S(o), and a functional S-allele, Sx, have previously been found to be retained in an F1 hybrid of a self-compatible commercial cultivar of Petunia hybrida. Pistil proteins associated with these two alleles have also been identified. Their amino-terminal sequences have been found to share a high degree of similarity with those of S-proteins characterized from self-incompatible solanaceous species. Here we report the isolation and sequencing of cDNAs encoding S(o)- and Sx-proteins. Their deduced amino acid sequences contain all the consensus primary structural features of S-proteins from self-incompatible solanaceous species. Both proteins also have ribonuclease activity. The implications of these findings are discussed in relation to the presumed function of the S-protein in the self-incompatibility interaction.
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PMID:Cloning and sequencing of cDNAs encoding two S proteins of a self-compatible cultivar of Petunia hybrida. 134 83

Hydrophobic interactions between the S-peptide and S-protein in the ribonuclease-S complex are probed using molecular dynamics simulations and free energy calculations. Three successive mutations at the buried position Met13 are simulated: Met----Leu, Leu----Ile, and Ile----Val, for which X-ray structures and experimental thermodynamic data are available. The calculations give theoretical estimates of the changes in binding free energies associated with these mutations. The calculated free energy differences are small (0-1.6 kcal/mol), in agreement with experiment. However the simulated structures deviate significantly from the experimental ones (mean deviation approximately 1.5-2 A), and a large uncertainty in the calculated free energies (1-2 kcal/mol) arises from the multiple minimum problem. Indeed, multiple conformations are available to the side chains around the mutation site, and the sampling of dihedral rotamer transitions is limited, despite long simulations. Fluctuations within each local minimum give rise to a small statistical error. However the uncertainty due to multiple conformations is much greater than the uncertainty due to random statistical errors. In our work, an artificial cancellation of errors arose because we studied conformations of the RNase complex and of the S-peptide that were very similar. In general, the criterion for a precise simulation is not merely to reduce the random statistical error, as has been suggested, but rather to sample all the important local minima along the mutation pathway, and to reduce the statistical error for each one. Our calculations suggest that the packing changes associated with the mutations are energetically small and localized, and largely cancel when the complex and the S-peptide are compared. Solvation of the methionine side chain partial charges in the S-peptide and the complex appear to be energetically equivalent, so that removing them (as in Met13----Leu, Ile, Val) does not affect binding. Enthalpy and entropy changes could not be estimated reliably.
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PMID:Thermodynamics of protein-peptide interactions in the ribonuclease-S system studied by molecular dynamics and free energy calculations. 139 Jun 51


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