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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Reversible unfolding of bovine chymotrypsinogen A in 2H2O either by heating at low pH or by exposure to 6 M guanidinium chloride results in the exchange of virtually all the nitrogen-bound hydrogens that give rise to low-field 1H NMR peaks, without significant exchange of the histidyl ring Cepsilon1 hydrogens. These preexchange procedures have enabled the resolution of two peaks, using 250-MHz correlation 1H NMR spectroscopy, that are attributed to the two histidyl residues of chymotrypsinogen A. Assignments of the Cepsilon1 hydrogen peaks to histidine-40 and -57 were based on comparison of the NMR titration curves of the native zymogen with those of the diisopropylphosphoryl derivative. Two histidyl Cepsilon1 H peaks were also resolved with solutions of preexchanged chymotrypsin Aalpha. The histidyl peaks of chymotrypsin Aalpha were assigned by comparison of NMR titration curves of the free enzyme with those of its complex with bovine pancreatic trypsin inhibitor (Kunitz). The NMR titration curves of histidine-57 in the zymogen and enzyme and histidine-40 in the zymogen exhibit two inflections; the additional inflections were assigned to interactions with neighboring carboxyl groups: aspartate-102 in the case of histidine-57 and aspartate-194 in the case of histidine-40 of the zymogen. In bovine chymotrypsinogen A in 2H2O at 31 degrees C, histidine-57 has a pK' of 7.3 and aspartate-102 a pK' of 1.4, and the histidine-40-aspartate-194 system exhibits inflections at pH 4.6 and 2.3. In bovine chymotrypsin Aalpha under the same conditions, the histidine-57-aspartate-102 system has pK' values of 6.1 and 2.8, and histidine-40 has a pK' of 7.2. The results suggest that the pK' of histidine-57 is higher than the pK' of aspartate-102 in both zymogen and enzyme. A significant difference exists in the structure and properties of the catalytic center between the zymogen and activated enzyme. In addition to the difference in pK' values, the chemical shift of histidine-57, which is highly abnormal in the zymogen (deshielded by 0.6 ppm), becomes normalized upon activation. These changes may explain part of the increase in the catalytic activity upon activation. The 1H NMR chemical shift of the Cepsilon1 H of histidine-57 in the chymotrypsin Aalpha-pancreatic trypsin inhibitor (Kunitz) complex is constant between pH 3 and 9 at a value similar to that of histidine-57 in the porcine trypsin-pancreatic trypsin inhibitor complex [Markley, J.L., and Porubcan, M. A. (1976), J. Mol. Biol. 102, 487--509], suggesting that the mechanisms of interaction are similar in the two complexes.
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PMID:Zymogen activation in serine proteinases. Proton magnetic resonance pH titration studies of the two histidines of bovine chymotrypsinogen A and chymotrypsin Aalpha. 3 98

Conformation in aqueous solution of adenosine-3',5'-cyclophosphate, 8-(beta-aminoethylamino) adenosine-3',5'-cyclophosphate, 8-(beta-oxiethylamino) adenosine-3',5'-cyclophosphate, 8-(carboxymethylamino) adenosine-3',5'-cyclophosphate and their non-cyclic analogs has been studied by NMR spectroscopy. The conformational situation in the model of dynamic equilibrium of sin- and anti-states has been described on the basis of spinlattice relaxation times and temperature dependences of chemical shifts. Adenosine-3',5'-cyclophosphate has been demonstrated to exist mainly in anti-conformation while 8-substituted analogs -- in sin-conformation. Equilibrium constants have been calculated for the compounds under study.
Mol Biol (Mosk)
PMID:[Nuclear magnetic resonance study of the conformation in nucleotides, oligonucleotides, and their analogs. I. Conformation of adenosine-3',5'-cyclic phosphate and its analogs in aqueous solutions]. 22 36

Four titrating histidine ring C2 and C4 proton resonances are observed in 220 MHz proton NMR spectra of human metmyoglobin as a function of pH. Values of ionization constants determined from the NMR titration data using an equation describing a simple proton association-dissociation equilibrium are curves (1) 6.6, (2) 7.0, (3) 5.8, and (4) 7.4. Four histidine residues have also been found to be solvent-accessible in human metmyoglobin by carboxymethylation studies (Harris, C.M., and Hill, R.L. (1969) J. Biol. Chem. 244, 2195-2203). Two of the titration curves (3 and 4) deviate significantly from the chemical shift values normally observed for histidine C2 proton resonances. Curve 3, with a low pKa, is shifted downfield at high values of pH and also exhibits a second minor inflection with a pKa value of 8.8. On the other hand, the high pKa curve, 4, is shifted upfield at all values of pH. The characteristics of the NMR titration curves with the lowest and highest pKa values (3 and4) are very similar to curves observed previously with sperm whale and horse metmyoglobins (Cohen, J.S., Hagenmaier, H., Pollard, H., and Schechter, A.N. (1972) J. Mol. Biol. 71, 513-519). These results indicate that the histidine residues from which these curves are derived have unusual and characteristic environments in this series of homologous proteins. The NMR spectra of all three metmyoglobins are changed extensively as a result of azide ion binding, indicating conformational changes affecting the environments of several imidazole side chains. The presence of azide ion causes a selective downfield chemical shift for the low pKa curve and a selective upfield chemical shift for the high pKa curve in all three proteins. Azide also abolishes the second inflection seen in the low pKa curve at high pH. In addition to these effects, the presence of azide ion permits the observation of two additional titrating proton resonances for all three metmyoglobins. Increasing the azide to protein ratio at several fixed values of pH yields results which show that a slow exchange process is occurring with each of the metmyoglobins. In the azide titration studies the maximum changes in the NMR spectra occurred at approximately equimolar concentrations. The NMR results for these proteins in the absence and presence of azide ion are related to x-ray crystallographic studies of sperm whale metmyoglobin and the known alkylation properties of the histidine residues. Tentative assignments of the titrating resonances observed are suggested.
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PMID:Nuclear magnetic resonance titration curves of histidine ring protons. Human metmyoglobin and the effects of azide on human, horse, and sperm whale metmyoglobins. 24 Aug 29

The high resolution 1H and 31P NMR spectra of the (dG-dC)8 duplex have been recorded in low- and high-salt solutions in order to evaluate the structural aspects of the salt-induced transition of oligo(dG-dC) in solution [Pohl, F. M. & Jovin, T. M. (1972) J. Mol. Biol. 67, 375-396]. The NMR data require that the (dG-dC)8 duplex in 4 M NaCl adopt an "alternating B-DNA" conformation for which the symmetry unit repeats every two base pairs. By contrast, the oligomer duplex in low-salt solution is of the regular B-DNA type in solution. The chemical shift parameters for oligo(dG-dC) in high-salt solution demonstrate that every other glycosidic torsion angle and phosphodiester linkage adopts a different conformation from that observed in regular B-DNA. We demonstrate further that the generation of the "alternating B-DNA" structure is facilitated by introduction of halogen atoms at the 5 position of pyrimidine and that this probably reflects the greater overlap of this position with adjacent base pairs in high salt solution. An "alternating B-DNA" model has recently been proposed for alternating deoxy purine-deoxy pyrimidine polynucleotides based on the x-ray structure of pdA-dT-dA-dT [Klug, A., Jack, A., Viswamitra, M.A., Kennard, O., Shakked, Z. & Steitz, T.A. (1979) J. Mol. Biol., in press].
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PMID:"Alternating B-DNA" conformation for the oligo(dG-dC) duplex in high-salt solution. 28 40

Optical and fluorescent characteristics of fluorescein covalently attached to 3'-end of tRNAFhe and X-nucleotide in the extra arm of several species of tRNA from E. coli have been studied. The probe is shown to be a sensitive factor indicating the conformational change of tRNA induced by Mg2+ and Na+ ions. By measuring the extent of energy transfer the distances between the fluorescent probe attached to 3'-terminus and X-nucleotide of tRNA and specific binding site of ethidium bromide on tRNA were determined to be 40.5 A and 32.5 A, respectively. The distances measured are in good agreement with the NMR spectroscopy data showing that the specific binding site for ethidium bromide on tRNA is localised near the sixth base pair of the acceptor stem.
Mol Biol (Mosk)
PMID:[Study of the structure of tRNA by the energy migration method using fluorescent labels covalently bound to specific tRNA loci]. 34 62

Rats of the Wistar/Af/Han/Mol/(Han 67) strain have previously been shown to respond in a variable way to phenobarbital treatment, as far as the induction of aldehyde dehydrogenase activity is concerned (Marselos 1976). This biochemical property is genetically determined and concerns the high-Km aldehyde dehydrogenase of the hepatic cytosol. In this study, administration of phenobarbital (1 mg/mo of drinking water, for 1 week) produces a uniform induction of aldehyde dehydrogenase in all rats, when measured with micromolar substrate concentration. The inducible low-Km enzyme of the cytosol is not genetically determined like the high-Km enzyme, and shows a wide specificity for aliphatic as well as for aromatic aldehydes. Despite the inducibility of the cytosolic enzymes, no alterations are found in the mitochondrial aldehyde dehydrogenase activities after phenobarbital treatment. The oxidation of D-glucuronolactone takes place only in the cytosol, and seems to be dependent on the low-Km aldehyde dehydrogenase. This is consistent with NMR studies, which showed that a very minimal amount of D-glucuronolactone is in aldehyde form under the measurement conditions usually applied. Further, the oxidation of D-glucuronolactone is also enhanced by phenobarbital in all rats without a genetic predisposition, and its dose-response curve is very similar to that of the low-Km aldehyde dehydrogenase.
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PMID:Inducible aldehyde dehydrogenases in the hepatic cytosol of the rat. 57 55

Natural abundance carbon-13 and proton NMR spectra of bovine chromaffin granules have been obtained and analyzed using computer simulation techniques. High resolution spectra show the presence of a fluid aqueous phase containing epinephrine, ATP and a random coil protein. The protein spectrum contains unusually intense resonances due to glutamic acid and proline and has been simulated satisfactorily using the known amino acid composition of chromogranin A. The lipid phase of chromaffin granules gives rise to intense, but very broad, resonances in the carbon-13 spectrum. Protons in the lipid phase are also observable as a very rapid component of the proton-free induction decay (T2 approximately equal to 15 microns). Linewidths of the carbon-13 spectra have been used to set upper limits on rotational correlation times and on the motional anisotropy in the aqueous phase. These limits show that the aqueous phase is a simple solution (not a gel) that is isotropic over regions much larger than solute dimensions. No gel transition is observed between -3 and 25 degrees C. The carbon-13 spectra are definitely inconsistent with a lipoprotein matrix model and chromaffin granules previously proposed by Helle and Serck-Hanssen ((1975) Mol. Cell, Biochem. 6, 127-146). Relative carbon-13 intensities of ATP and epinephrine are not consistent with the known 1 : 4 mol ratio of these components. This fact suggests that epinephrine and ATP are not directly complexed in intact chromaffin granules.
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PMID:Analysis of the carbon-13 and proton NMR spectra of bovine chromaffin granules. 84 74

The methods which have been used for the observation and assignment of resonances in the NMR spectra of proteins are reviewed. One such method, the selective deuteration of the aromatic protons of tryptophyl residues, is studied by NMR spectroscopy in model compounds in this paper, and in proteins in the following paper. On the basis of a reassignment of the PMR spectrum of the aromatic protons of L-tryptophan, the relative rates of H-D exchange in deutero-trifluoracetic acid (d-TFA) are H-2 greater than H-5 greater than H-6 greater than H-4 approximately H-7. The energies of activation for the first order exchange of both the H-2 and H-5 protons is 12 k.cal.mol-1. The rate constant for exchange of the H-2 protons of tryptophyl residues in peptides is much greater than in the amino acid itself and 5-10 times that for exchange of the H-5 protons. This suggests that the method can be used to label tryptophyl residues in proteins rapidly and specifically.
Mol Cell Biochem 1976 Aug 30
PMID:Kinetics of hydrogen-deuterium exchange of tryptophan and tryptophan peptides in deutero-trifluoroacetic acid using proton magenetic resonance spectroscopy. 95 13

NMR spectra of the downfield region of normal adult hemoglobin are reported as a function of oxygenation and temperature. Spectra were run in D2O at pD 7.4. A specially made NMR tube insert allowed precise measurement of the degree of oxygenation and of methemoglobin formation before and after taking the NMR spectrum. Plots of the estimated intensity of the most downfield prominent NMR peak, identified as arising from a deoxy-beta subunit by Davis et al. ((1971) J. Mol. Biol. 60, 101-111), versus the average degree of oxygenation y, measured optically, yield a nearly straight line within experimental error, for samples stripped of organic phosphates and for samples containing 2,3-diphosphoglycerate or inositol hexaphosphate. Intensities of peaks further upfield than this peak, previously attributed to deoxy-alpha subunits, are difficult to measure directly especially for samples containing inositol hexaphosphate. The latter samples show broadening in these alpha peaks as the degree of oxygenation increases. This extra broadening appears to increase with temperature. Linearity of the beta peak intensity with oxygenation is expected if there is no large oxygen affinity difference between alpha and beta subunits. However, the cooperativity of binding, and inaccuracy of the data, make it impossible to make accurate estimates of affinity differences.
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PMID:NMR study of relative oxygen binding to the alpha and beta subunits of human adult hemoglobin. 99 7

A differential method is described for measuring dielectric constants and losses in aqueous protein solutions at millimetrerange wavelengths. Employment of the method allows to improve the accuracy of determining the degree of hydratation. A method has also been suggested for taking into account the contribution of ions to the dielectric constant of solutions. The differential method was used to study hydratation of nine globular proteins. The data obtained are compared with the corresponding values provided by other experimental techniques and with theoretical predictions based on some models of hydratation. Good agreement is obtained with results provided by the isopiestic and NMR techniques. The discrepancy shown for hemoglobin is discussed in the paper. As has been shown, the dielectric method registers a monomolecular surface layer of water only. With pH varying between 4.0 and 3.2, a significant increase is observed in the hydratation of serum albumin. Presumably, this effect is connected with a N--F conformational transition.
Mol Biol (Mosk)
PMID:[Globular protein hydration by a differential dielectrometric method]. 105 42


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