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)

It was found, in cell-free assays, that the Man8GlcNAc2 and Man7GlcNAc2 isomers having the mannose unit to which the glucose is added were glucosylated by the rat liver glucosyltransferase at 50 and 15%, respectively, of the rate of Man9GlcNAc2 glucosylation. This indicates that processing by endoplasmic reticulum mannosidases decreases the extent of glycoprotein glucosylation. All five different glycoproteins tested (bovine and porcine thyroglobulins, phytohemagglutinin, soybean agglutinin, and bovine pancreas ribonuclease B) were found to be poorly glucosylated or not glucosylated unless they were subjected to treatments that modified their native conformations. The effect of denaturation was not to expose the oligosaccharides but to make protein determinants, required for enzymatic activity, accessible to the glucosyltransferase because (a) cleavage of denatured glycoproteins by unspecific (Pronase) or specific (trypsin) proteases abolished their glucose acceptor capacities almost completely except when the tryptic peptides were held together by disulfide bonds and (b) high mannose oligosaccharides in native glycoproteins, although poorly glucosylated or not glucosylated, were accessible to macromolecular probes as concanavalin A-Sepharose, endo-beta-N-acetylglucosaminidase H, and jack bean alpha-mannosidase. In addition, denatured, endo-beta-N-acetylglucosaminidase H deglycosylated glycoproteins were found to be potent inhibitors of the glucosylation of denatured glycoproteins. It is suggested that in vivo only unfolded, partially folded, and malfolded glycoproteins are glucosylated and that glucosylation stops upon adoption of the correct conformation, a process that hides the protein determinants (possibly hydrophobic amino acids) from the glucosyltransferase.
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PMID:Recognition of the oligosaccharide and protein moieties of glycoproteins by the UDP-Glc:glycoprotein glucosyltransferase. 153 Oct 24

The characterization of high-mannose-type N-glycosylation by capillary electrophoresis-electrospray mass spectrometry (CE-ESI MS) was described. In addition to the use of a cationic noncovalent capillary coating, strong acidic buffer, and charge reversal to increase the glycoform resolving power, N-glycosidase F (PNGase F) combined with a basic protease and alpha-mannosidase combined with an acidic protease were used to analyze the high-mannose-type N-glycosylation in ribonuclease B (RNase B) and in a novel C-type lectin from the venom of Trimeresurus stejnegeri (TSL). The structures of oligosaccharide, glycosylation sites, and glycoform distributions were determined simultaneously, and the differential oxidation of Met residues in glycopeptides obtained from TSL protease digestion was also characterized successfully by CE-MS/MS. The results showed that the oligosaccharide attached to RNase B has a structure of GlcNAc2Man5 approximately 9, and that attached to TSL has a structure of GlcNAc2Min5 approximately 8. The glycoform distributions in these glycoproteins are quite different, with the GlcNAc2Man5 type predominant in RNase B, and the GlcNAc2Man8 type, in TSL This method may be useful not only for the characterization of glycosylation sites and glycan structures, but also for the determination of the relative abundance of individual glycoforms.
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PMID:Capillary electrophoresis-electrospray mass spectrometry for the characterization of high-mannose-type N-glycosylation and differential oxidation in glycoproteins by charge reversal and protease/glycosidase digestion. 1179 56

Liquid chromatography-electrospray mass spectrometry was utilized to analyze peptide mapping of a glycoprotein ribonuclease B to obtain its primary structure. The glycosylated site was determined by comparison of peptide mapping before and after glycanase treatment.Tandem MS(MS/MS)was performed to analyze the structure of N-linked glycan and deglycosylated peptide. The nature of glycan was determined to be of highmannose type by mass spectrometry after the treatment with alpha-mannosidase. In addition the relative abundance of heterogeneous glycopeptides was quantified. This method is rapid and sensitive for the characterization of glycoproteins with N-linked glycan.
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PMID:Analysis of Peptide Mapping Glycosylated Site and Glycan Structure in Ribonuclease B by Liquid Chromatography-Electrospray Mass Spectrometry. 1211 Sep 38

We present evidence that protein bodies constitute the principal lytic compartment in storage parenchyma cells of mung bean cotyledons and propose that they play a role in cellular autophagy. We developed a method to isolate protein bodies by incubating tissue slices with cell wall-degrading enzymes and fractionating the cellular organelles on a Ficoll gradient. About 75-80% of the protein bodies present in the protoplasts were recovered intact in a band at the 5/25% Ficoll interface. This band contained a similar proportion of the cellular alpha-mannosidase, N-acetyl-beta-glucosaminidase, ribonuclease, acid phosphatase, phosphodiesterase, and phospholipase D. beta-Amylase was present in the cells but not in the protein bodies. Ultrastructural observations showed that on the 3rd day of seedling growth protein bodies contain small vesicles (0.3-1.0 mum) with a cytoplasmic content (ribosomes, membrane vesicles, mitochondria). Later in seedling growth these vesicles appeared empty. We believe that these are autophagic vesicles resulting from invaginations of the protein body membrane and that their cytoplasmic contents are digested by the acid hydrolases present in the protein bodies.
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PMID:Protein bodies of mung bean cotyledons as autophagic organelles. 1659 58

Recombinant glycoproteins present unique challenges to biopharmaceutical development, especially when efficacy is affected by glycosylation. In these cases, optimizing the protein's glycosylation is necessary, but difficult, since the glycan structures cannot be genetically encoded, and glycosylation in nonhuman cell lines can be very different from human glycosylation profiles. We are exploring a potential solution to this problem by designing enzymatic glycan optimization methods to produce proteins with useful glycan compositions. To demonstrate viability of this new approach to generating glycoprotein-based pharmaceuticals, the N-linked glycans of a model glycoprotein, ribonuclease B (RNase B), were modified using an alpha-mannosidase to produce a new glycoprotein with different glycan structures. The secondary structure of the native and modified glycoproteins was retained, as monitored using circular dichroism. An assay was also developed using an RNA substrate to verify that RNase B had indeed retained its function after being subjected to the necessary glycan modification conditions. This is the first study that verifies both activity and secondary structure of a glycoprotein after enzymatic glycan trimming for use in biopharmaceutical development methods. The evidence of preserved structure and function for a modified glycoprotein indicates that extracellular enzymatic modification methods could be implemented in producing designer glycoproteins.
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PMID:Development of fully functional proteins with novel glycosylation via enzymatic glycan trimming. 1911 44