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)

Recent work has shown that, with synthetic analogues of C-peptide (residues 1-13 of ribonuclease A), the stability of the peptide helix in H2O depends strongly on the charge on the N-terminal residue. We have asked whether, in semisynthetic ribonuclease S reconstituted from S-protein plus an analogue of S-peptide (1-15), the stability of the peptide helix is correlated with the Tm of the reconstituted ribonuclease S. Six peptides have been made, which contain Glu9----Leu, a blocked alpha-COO- group (-CONH2), and either Gln11 or Glu11. The N-terminal residue has been varied; its charge varies from +2 (Lys) to -1 (succinyl-Ala). We have measured the stability of the peptide helix, the affinity of the peptide for S-protein (by C.D. titration), and the thermal stability of the reconstituted ribonuclease S. All six peptide analogues show strongly enhanced helix formation compared to either S-peptide (1-15) or (1-19), and the helix content increases as the charge on the N-terminal residue changes from +2 to -1. All six peptides show increased affinity for S-protein compared to S-peptide (1-19), and all six reconstituted ribonucleases S show an increase in Tm compared to the protein with S-peptide (1-19). The Tm increases as the charge on residue 1 changes from +2 to -1. The largest increment in Tm is 6 degrees. The results suggest that the stability of a protein can be increased by enhancing the stability of its secondary structure.
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PMID:The design and production of semisynthetic ribonucleases with increased thermostability by incorporation of S-peptide analogues with enhanced helical stability. 344 49

Gabonase, an enzyme which acts on fibrinogen and factor XIII in uniquely thrombin-like ways, was purified to electrophoretic homogeneity from the venom of Bitis gabonica. On sodium dodecyl sulfate-polyacrylamide electrophoresis, the reduced protein behaved as a single chain with Mr = 30,600. The enzyme contains 20.6% carbohydrate, no free sulfhydryl groups and hence, from amino acid analysis, five disulfide bonds. Its extinction coefficient (E1%1cm) at 280 nm is 9.6. Its pI is 5.3. Gabonase has an active serine residue, is inactivated by phenylmethanesulfonyl fluoride, and has an active histidine which reacts with the chloromethyl ketone of tosyl-L-lysine. Its NH2-terminal amino acid sequence (Val-Val-Gly-Gly-Ala-Glu-Cys-Lys-Ile-Asp-Gly-His-Arg-Cys-Leu-Ala-Leu-Leu -Tyr-) is homologous to the B chain of thrombin. The activity of the enzyme is stabilized by calcium ion. It exhibits strong N alpha-p-tosyl-L-arginine methyl esterase activity, hydrolyzes tripeptide nitroanilide derivatives weakly or not at all, and cleaves no peptide bonds in insulin, glucagon, or the S peptide of ribonuclease. Gabonase clots fibrinogen with a specific activity of 45 NIH thrombin-equivalent units/mg, releasing both fibrinopeptides A and B and showing substrate inhibition at fibrinogen concentrations of 3 mg/ml or greater. The enzyme also activates factor XIII. It is not inactivated by either heparin or hirudin.
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PMID:Thrombin-like enzyme from the venom of Bitis gabonica. Purification, properties, and coagulant actions. 352 80

The 1H-n.m.r. spectra (360 MHz) of 12-(beta-(3-pyridyl)-L-Ala) ribonuclease S-peptide (1-14), a tetradecapeptide incorporating (beta-3-pyridyl-L-Ala) instead of His at position 12, have been assigned. The shift vs. temperature dependence has been analyzed at three different pD's in terms of a two-state helix (3-13) in equilibrium coil equilibrium, and the corresponding values for the thermodynamic quantities delta H degrees and delta S degrees determined. Helix populations at 0 degrees C have been measured as a function of pD, showing their dependence on two apparent pKa's at approximately 3.3 and 5.5, with a maximum at pD approximately 4.2. All the obtained results show that the new peptide has very similar folding properties to those shown by S-peptide and particularly to those of C-peptide. The 3-13 helix formed is stabilized by two interactions: a salt-bridge Glu 2-...Arg 10+ and a partial stacking between the aromatic rings of residues Phe 8 and His 12. Calculations involving ring current shifts and potential energies validate the possible existence of this latter interaction, which must present a local geometry defined by chi 81 180 degrees, chi 82 100 degrees, chi 121-60 and chi 122 80.
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PMID:1H-n.m.r. study of the folding of ribonuclease 12-(beta-(3-pyridyl)-L-Ala) S-peptide (1-14). 357 Jun 61

A single stained band containing approximately 5 micrograms of protein was cut out of a polyacrylamide gel and subjected to hydrolysis together with the gel. The hydrolysate was subsequently analyzed for its amino acid content by high-performance liquid chromatography and postlabeling with o-phthalaldehyde. Bovine serum albumin, ribonuclease B, ovalbumin, pepsin, and chymotrypsinogen A were analyzed by this method, and their amino acid compositions were found to be in good agreement with the reported values. By this method, it is possible to quantitate 16 amino acids: Asx, Thr, Ser, Glx, Pro, Cys, Gly, Ala, Val, Ile, Leu, Tyr, Phe, His, Lys, and Arg. Thioglycolic acid is effective protection against the decomposition of Tyr, Cys, and Met; however, the recovery of Met is inconsistent. This method might be very helpful for the amino acid analysis of proteins of multicomponent systems, especially, those which can be resolved only by polyacrylamide gel electrophoresis.
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PMID:Amino acid analysis by high-performance liquid chromatography of a single stained protein band from a polyacrylamide gel. 357 64

A base non-specific and adenylic acid preferential ribonuclease from Aspergillus saitoi (RNase M) was modified by [14C]iodoacetic acid. RNase M was inactivated with concomitant incorporation of about 1 mol equivalent of carboxymethyl group. Carboxymethylated RNase M (CM RNase M) thus obtained was reduced and carboxymethylated (RCM CM RNase M). From tryptic and chymotryptic digests of RCM CM RNase M, two carboxymethylated histidine-containing peptides labeled with radioactivity were isolated. The amino acid sequences of these two peptides were determined to be Thr-Ile-His-Gly-Leu-Trp-Pro-Asp-Asn-Cys-Asp-Gly-Ser-Tyr... and His-Gly-Thr-Cys-Ile-Asn-Thr-Ile-Asp-Pro-Ser-Cys-Tyr-Pro-Asp-Asp-Tyr-Ala. .... The distribution of the radioactivity on the former and latter peptides was 43% and 57%, respectively. The results indicated that two histidine residues are involved in the active site of RNase M, and the modification of either one of the two histidine residues inactivates RNase M. The CD spectrum of carboxymethylated RNase M indicated that some tryptophan residue(s) with a CD band at 287 nm is in the proximity of the active site histidine residues of RNase M.
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PMID:Site of alkylation of the major ribonuclease from Aspergillus saitoi with iodoacetate. 371 Oct 38

The refolding of urea-denatured ribonuclease A was measured at 0.31-3.1 mol . l-1 urea in the presence of various concentrations of peptidyl-prolyl cis-trans isomerase isolated from pig kidney. The rate of the slow CT-phase in the refolding reaction was found to be sensitive to this enzyme. A rate enhancement proportional to the isomerase activity has been observed. The activity of the enzyme was assayed with Glt-Ala-Ala-Pro-Phe-4-nitroanilide. The catalytic activity of the isomerase against unfolded ribonuclease is suppressed after preincubation of the enzyme with 0.001 mol . l-1 Cu2+, 0.01 mol . l-1 H+ and by heat inactivation. The results indicate the involvement of the cis/trans interconversion of proline peptide bonds during the refolding of ribonuclease A.
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PMID:The refolding of urea-denatured ribonuclease A is catalyzed by peptidyl-prolyl cis-trans isomerase. 388 50

In order to examine the effect of a defined enantiomeric sequence on protein structure, the all-D model ribonuclease S-peptide, H-Ala-Glu-Ala4-Lys-Phe-Ala-Arg-Ala-His-Met-Ala2-OH, has been synthesized by the solid phase method. The all-L peptide has been synthesized previously and shown to possess 36% of ribonuclease S activity when added to ribonuclease S-protein (Komoriya, A. & Chaiken, I.M. (1982) J. Biol. Chem 257, 2599-2604). The synthetic D-peptide was purified by gel filtration and semipreparative reverse phase HPLC. Amino acid composition of the synthetic peptide was in agreement with theory and gas chromatographic analysis showed that no significant racemization had occurred during synthesis. Circular dichroism (CD) studies of the D-peptide showed a peak of positive ellipticity in the 220-230 nm region, whereas a negative ellipticity peak for the L-peptide was observed. The effects of temperature and trifluoroethanol on the far-ultraviolet CD spectra of D- and L-peptides were similar but of opposite sign, confirming the expectation that the D-peptide has the propensity to form an alpha-helical structure which is enantiomeric with respect to that formed by the L-peptide. In the presence of S-protein, the L-peptide showed hydrolytic activity against the substrate cytidine-2':3'-monophosphate, whereas the D-peptide was inactive. Addition of the D-peptide to mixtures of L-peptide and S-protein did not lead to inhibition of enzymatic activity. These results indicate lack of binding of D-peptide to S-protein to produce either an active or inactive species.
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PMID:Synthesis and properties of an all-D model ribonuclease S-peptide. 399 53

Snapping turtle (Chelydra serpentina) ribonuclease was isolated from pancreatic tissue. Turtle ribonuclease binds much more weakly to the affinity chromatography matrix used than mammalian ribonucleases. The amino acid sequence was determined from overlapping peptides obtained from three different digests. The N-terminal amino acid sequence of the protein determined by others [E. A. Barnard, M. S. Cohen, M. H. Gold J.-K. Kim (1972) Nature (Lond.) 240, 395-398] and homology were used as additional evidence for several overlaps. The polypeptide chain consists of 119 amino acid residues. Compared to most ribonucleases the N-terminal residue, three residues in the loop near residue 71 and two residues in the loop near residue 114 are deleted, and there is one additional residue in the loop near residue 23. The half-cystines at positions 65 and 72, which form a disulfide bond in mammalian ribonucleases, are not present in turtle ribonuclease. Turtle ribonuclease differs from bovine ribonuclease at 70 of the 118 positions where both proteins have amino acid residues. Turtle ribonuclease contains no carbohydrate, although the enzyme possesses a recognition site for carbohydrate attachment in the sequence Asn-Ala-Ser (positions 76-78).
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PMID:The amino acid sequence of snapping turtle (Chelydra serpentina) ribonuclease. 407 78

1. The alanyl-s-RNA synthetase of tomato roots has been purified by ammonium sulphate precipitation, adsorption on calcium phosphate gel and DEAE-cellulose chromatography and its properties have been investigated. 2. Enzyme activity was measured by using the hydroxamate assay, the [(32)P]pyrophosphate-ATP-exchange assay and the [(14)C]alanyl-s-RNA assay. The purified enzyme was specific for l-alanine and was activated by Mg(2+) ions and to a smaller extent by Co(2+) and Mn(2+) ions. It was free from adenosine triphosphatase, pyrophosphatase and ribonuclease, and possessed a specific activity comparable with that of the most highly purified aminoacyl-s-RNA synthetases from animal and microbial systems. 3. The properties of the purified enzyme were similar in many respects to most other highly purified aminoacyl-s-RNA synthetases. It differed, however, in that the pH optimum of the hydroxamate assay was almost the same as that of the pyrophosphate-ATP-exchange assay and in requiring a high concentration of l-alanine for maximum activity (100mumoles/ml.). 4. The purified enzyme was not absolutely specific for tomato-root s-RNA; slight activity was also observed with yeast s-RNA. 5. The properties of this enzyme are fully consistent with the suggestion that the enzymic formation of alanyl-s-RNA proceeds via the intermediate formation of alanyl acyl-adenylate with the elimination of pyrophosphate from ATP. It remains to be shown the extent to which alanyl-s-RNA participates further in subsequent stages of protein synthesis in plants.
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PMID:The purification and properties of the alanyl-transfer ribonucleic acid synthetase of tomato roots. 428 91

Chick embryo tRNA, prepared by a simple large-scale method, was fractionated on three different ion-exchange columns. In all cases simple chromatographic patterns for various tRNA species were observed, indicating the presence of only a few major species of tRNA for each amino acid. By repeated chromatography one species of alanine tRNA was purified to approx. 80% purity. T(1) ribonuclease digest of this purified tRNA gave a simple chromatographic pattern. Because of the simplicity of the method of preparation of tRNA from this readily available source and the presence of only a few species of tRNA for each amino acid, chick embryo is suited for the study of tRNA and its various functions in higher systems.
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PMID:Studies on the fractionation of total transfer ribonucleic acid and purification of an alanine transfer ribonucleic acid from chick embryo. 446 55


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