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)

The binding of immunogenic peptides to class II major histocompatibility molecules was examined at various pH values. We studied binding of peptides containing residues 52-61 from hen egg lysozyme (HEL) to I-Ak on fixed peritoneal macrophages or to solubilized affinity-purified I-Ak. Optimum binding occurred at pH 5.5-6.0 with accelerated kinetics relative to pH 7.4; equilibrium binding was also higher at pH 5.5-6.0 than at 7.4. Similar enhancement at pH 5-6 was observed for the binding of hemoglobin-(64-76) to I-Ek and of ribonuclease-(41-61) to I-Ak. In contrast, the binding of HEL-(34-45) to I-Ak was minimally enhanced at acid pH. Dissociation of cell-associated or purified peptide-I-Ak complexes was minimal between pH 5.5 and 7.4, with increased dissociation only at or below pH 4.0 [HEL-(46-61)] or pH 5.0 [HEL-(34-45)]. Thus, optimum peptide binding occurs at pH values similar to the endosomal environment, where the complexes appear to be formed during antigen processing. In addition, we examined the effect of a number of polysaccharides on the binding of peptide to I-Ak. None of these competed with the HEL peptide 125I-labeled YE52-61 for binding to I-Ak. [3H]Dextran also failed to bind purified I-Ak. Polysaccharides do not appear to bind to class II major histocompatibility complex molecules, which explains the T-cell independence of polysaccharide antigens.
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PMID:Effects of pH and polysaccharides on peptide binding to class II major histocompatibility complex molecules. 201 83

Dissociation of RNA and DNA from Escherichia coli RNA polymerase in transcription complexes prepared with enzyme molecules located within and near a rho-dependent transcription termination region on bacteriophage T7 D111 DNA has been studied using a membrane filter-binding assay. Rho protein with ATP present mediated rapid (half-time approximately 27 s) simultaneous dissociation of about 50% of both RNA and DNA. RNA molecules were preferentially released from enzyme molecules located within the termination region. Rapid release of RNA and DNA depended on a nucleoside triphosphate but did not depend on sigma factor. Pretreatment of complexes with ribonuclease prevented dissociation of DNA. Nearly simultaneous dissociation of both RNA and DNA was also detected after a lag of 3 min when the isolated transcription complexes were incubated with all four ribonucleoside triphosphates in the absence of rho factor. In this case, release presumably occurred at the rho-independent termination site that is 5990 nucleotides downstream from the A1 promoter. Thus, the dissociation of DNA from RNA polymerase at rho-dependent and rho-independent transcription termination sites is coupled with or occurs spontaneously soon after the release of transcripts at both sites.
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PMID:Transcription termination factor rho mediates simultaneous release of RNA transcripts and DNA template from complexes with Escherichia coli RNA polymerase. 388 62

Rat uterine and anterior pituitary microsomes each contain a population of specific estrogen-binding sites. Saturation binding of estradiol is demonstrable, with an affinity similar to that of the cytosol estrogen receptor (Ka = 1-2 X 10(10) M-1). Dissociation rate kinetic determinations, however, revealed that estrogen-microsomal complexes are 4 times as stable as cytosol estrogen-receptor complexes. Sedimentation properties in sucrose gradients were salt-dependent, yielding values of 10S in KCl-free buffer and 5.5S in the presence of 0.4 M KCl. The concentration of microsomal sites varies in proportion to the level of cytosol estrogen receptor, such that microsomal binding constitutes a consistent 20% of the total extranuclear binding capacity. Binding is sensitive to pronase, but not to ribonuclease or deoxyribonuclease; steroidal specificity differs from cytosol receptor only with respect to a greater extent of competition by progesterone. Microsomal binding sites are readily extractable with KCl-free hypotonic buffer or with 0.4 M KCl, but are resistant to extraction by 0.15 M KCl. The presence of estradiol lends stability to the microsomal binding sites, while high salt has a deleterious effect on their longevity. After exhaustive extraction of binding sites, microsomes are capable of accepting cytosol estradiol-receptor complexes to a level corresponding to the concentration of depleted binding sites; microsomes from nontarget tissue do not manifest such capability. However, the original microsomal estrogen-binding sites are not simply cytosol receptor contaminants, as evidenced by the observations that the microsomal binding site concentration is independent of the volume of tissue homogenate (indicating that a trapping phenomenon is not operative) and that nonextracted microsomes are not potential acceptor sites for cytosol estradiol-receptor complexes. In considering total cellular dynamics of estrogen and estrogen receptor turnover, it thus becomes important to explore the role of the microsomal compartment, since it functions as a repository of specific estrogen-binding sites and may have significant acceptor capability for the cytosol estrogen-receptor complex.
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PMID:Specific binding of estrogen and estrogen-receptor complex by microsomes from estrogen-responsive tissues of the rat. 402 80

The binding efficiency of high affinity monoclonal antifluorescyl antibody 4.4-20 with the homologous ligand situated in different protein environments has been investigated to quantitate the effect of non-active site secondary factors. To synthesize monofluoresceinated proteins, fluorescein 5-isothiocyanate was reacted with a 100-fold molar excess of ribonuclease, lysozyme, lactalbumin and bovine serum albumin. Absorption and emission spectra, as well as fluorescence life-time measurements which yielded discrete components and proteolytic studies suggested that fluorescein was conjugated to a specific lysine residue consistent with a non-random distribution of lysines within each protein population. The derivatized residue was probably a surface moiety based on accessibility analyses with iodide as a dynamic quencher. Dissociation rate analyses indicated that the relative release time of 4.4-20 with each monofluoresceinated protein was Fl-RNAse > or = Fl-lyso > or = FDS > Fl-lact > or = Fl-BSA which correlated with changes in free energy of binding. Relative fluorescence quenching measurements of the fluorescein moiety indicated that 4.4-20 showed decreasing quenching in the order FDS > Fl-RNAse > Fl-lyso > or = Fl-lact > Fl-BSA. Because spectral data indicated that fluorescein was conjugated to a specific residue or a non-random distribution of residues in each protein population, the results represented the effect of a single distinct environment or a weighted average of different microenvironments. Results have been interpreted within the theoretical framework of a dynamic antibody model involving conformer selection and the relative effects of primary and secondary interactions.
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PMID:Effects of secondary forces on the primary antibody-ligand interaction. 855 47

The contribution of RNA degradation to the posttranscriptional control of gene expression confers on it a fundamental role in all biological processes. Ribonucleases (RNases) are essential enzymes that process and degrade RNA and constitute one of the main groups of factors that determine RNA levels in the cells. RNase II is a ubiquitous, highly processive hydrolytic exoribonuclease that plays an important role in RNA metabolism. This ribonuclease can act independently or as a component of the exosome, an essential RNA-degrading multiprotein complex. In this chapter, we explain the general procedures normally used for the characterization of ribonucleases, using as an example a study performed with Escherichia coli RNase II. We present the overexpression and purification of RNase II recombinant enzyme and of a large set of RNase II truncations. We also describe several methods that can be used for biochemically characterizing the exoribonucleolytic activity and studying RNA binding in vitro. Dissociation constants were determined by electrophoretic mobility shift assay (EMSA), surface plasmon resonance (SPR), and filter binding assays using different single- or double-stranded RNA substrates. We discuss the synergies among the biochemical analyses and the structural studies. These methods will be very useful for the study of other ribonucleases.
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PMID:Characterizing ribonucleases in vitro examples of synergies between biochemical and structural analysis. 1916 42

As one of the most prevalent and complex post-translational modifications in biological systems, proteins glycosylation has drawn considerable attention in recent decades. Dissociation of the carbohydrates from glycoproteins may be the prerequisite step of glycomics experiments, which commonly performed by specific proteolysis. In this study, an alternative strategy was reported with nonspecific proteolysis in coupling with co-derivatization of TMPP-Ac and methylamidation for glycan moieties analysis by MALDI-MS. With the co-derivatization, a permanent positive charge was introduced to the Asn-glycans and the carboxylic groups were neutralized by methylamidation simultaneously. As a result, approximately 20 and 50-fold enhancement in the detection sensitivity was achieved for asialo-Asn and disialo-Asn respectively in comparison to their native counterparts. Ultimately, this developed strategy was successfully validated using three model glycoproteins, including ribonuclease B, ovalbumin and transferrin.
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PMID:Matrix-assisted laser desorption/ionization mass spectrometry analysis of glycans with co-derivatization of asparaginyl-oligosaccharides. 2648 93