Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The sialidase from Salmonella typhimurium LT2 was characterized by using photoaffinity-labelling techniques. The well-known sialidase inhibitor 5-acetamido-2,6-anhydro-3,5-dideoxy-D-glycero-D-galacto-non- 2-enonic acid (Neu5Ac2en) was modified to contain an amino group at C-9, which permitted the incorporation of 4-azidosalicylic acid in amide linkage at this position. Labelling of the purified protein with the radioactive (125I) photoprobe was determined to be highly specific for a region within the active-site cavity. This conclusion was based on the observation that the competitive inhibitor Neu5Ac2en in the photolysis mixture prevented labelling of the protein. In contrast, compounds with structural and chemical features similar to the probe and Neu5Ac2en, but which were not competitive enzyme inhibitors, did not affect the photolabelling of the protein. The peptide interacting with the probe was identified by CNBr treatment of the labelled protein, followed by N-terminal sequence analysis. Inspection of the primary structure of the protein, predicted from the cloned structural gene for the sialidase [Hoyer, Hamilton, Steenbergen & Vimr (1992) Mol. Microbiol. 6, 873-884] revealed that the label was incorporated into a 9.6 kDa fragment situated within the terminal third of the molecule near the C-terminal end. Secondary-structural predictions using the Garnier-Robson algorithm [Garnier, Osguthorpe & Robson (1978) J. Mol. Biol. 120, 97-120] of the labelled peptide revealed a structural similarity to the active site of influenza-A- and Sendai-HN-virus sialidases with a repetitive series of alternating beta-sheets connected with loops.
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PMID:Photolabelling of Salmonella typhimurium LT2 sialidase. Identification of a peptide with a predicted structural similarity to the active sites of influenza-virus sialidases. 129 92

A genomic clone, pTt21, containing DNA apparently transcribed specifically in Trypanosoma cruzi trypomastigotes, was obtained by differentially screening a genomic library with trypomastigote and epimastigote cDNA. This 3444-bp clone contained open reading frames at each end, separated by a 1.8-kb non-coding region. The translated polypeptide from the 3' open reading frame (ORF2) of 1037 bp had 25-30% identity with 5 recently published T. cruzi gp85/sialidase sequences, and 20-25% identity with bacterial sialidases. Rabbit antiserum raised against an Escherichia coli fusion protein derived from the 5' open reading frame (ORF1) identified a surface antigen of 160 kDa, specifically expressed in trypomastigotes. A probe containing the first 211 bp from ORF1 was used to obtain a complete copy (c1821) of a gene that was closely related to ORF1, and encoded another member of the gp85/sialidase family. c1821 encodes a protein of 897 amino acids, but assignment of the N-terminus of the polypeptide was not possible. The 5'-most start codon is an unfavourable context to act as a translation initiator, it does not align with the initiator methionines of other gp85/sialidase sequences, nor is it followed by a signal peptide sequence characteristically found in other gp85/sialidase sequences. Although homology with the 5' ends of other gp85/sialidase sequences decays towards the 5' end of c1821, alignment of c1821 with 4 other gp85/sialidases indicated that the coding sequence should extend upstream at least 160 amino acids. In this region of c1821 there are multiple stop codons in each frame. The presence of the stop codons, the alignment data and our inability to amplify reverse transcribed mRNA using four internal primers, suggest that c1821 may not be present as a mature mRNA and is a pseudogene. Comparison of the apparently non-repetitive 3' coding domain of c1821 with the corresponding repetitive domains of two other members of the gp85/sialidase family revealed a high degree of similarity in nucleotide but not in amino acid sequence, and c1821 may thus represent an evolutionary intermediate between sub-families of the gp85/sialidase superfamily.
Mol Biochem Parasitol 1992 Nov
PMID:Sequence homology and absence of mRNA defines a possible pseudogene member of the Trypanosoma cruzi gp85/sialidase multigene family. 147 90

A membrane-bound sialidase (EC 3.2.1.18) was found in procyclic trypomastigotes of Trypanosoma brucei. The mammalian stage bloodstream form, however, displayed no sialidase activity. This sialidase is an integral surface protein, linked to the membrane via a glycosylphosphatidylinositol anchor. After osmotic lysis and solubilization with Triton CF-54, the enzyme was purified 1900-fold by gel filtration and ion exchange chromatography. Its size, as determined by conventional and high-performance liquid gel chromatography, is 67 kDa. The sialidase is active over a broad pH and temperature range with optima at pH 6.9 and 35 degrees C, respectively. No loss of activity is observed after 4 freeze-thaw cycles. T. brucei sialidase activity is inhibited by N-(4-nitrophenyl)oxamic acid and 2-deoxy-2,3-didehydro-N-acetylneuraminic acid, the latter, however, being less effective. N-Acetylneuraminic acid shows no inhibitory effect, whereas a variety of metal ions are potent inhibitors. The sialidase is activated by di- and tricarboxylic acids, but inhibited by chloride. Relative hydrolysis rates of various sialic acid-containing compounds reveal that de-O-acetylated bovine submandibular gland mucin is the preferred substrate and that alpha(2-3)-linkages are hydrolyzed faster than alpha(2-6)-linkages.
Mol Biochem Parasitol 1992 Aug
PMID:Purification and characterization of a novel sialidase found in procyclic culture forms of Trypanosoma brucei. 151 30

Sialic acid on human erythrocytes is involved in invasion by the human malaria parasite, Plasmodium falciparum. Mouse erythrocytes were used as a reagent to explore the question of whether erythrocyte sialic acid functions as a nonspecific negative charge or whether the sialic acid is a necessary structural part of the receptor for merozoites. Human erythrocytes contain N-acetylneuraminic acid (Neu5Ac), whereas mouse erythrocytes, which are also invaded by P. falciparum merozoites, contain 9-O-acetyl-N-acetylneuraminic acid (Neu5,9Ac2) and N-glycoloylneuraminic acid (Neu5Gc), in addition to Neu5Ac. We compared the effects of sialidase and influenza C virus esterase treatments of mouse erythrocytes on invasion and the binding of a 175-kDa P. falciparum protein (EBA-175), a sialic acid-dependent malaria ligand implicated in the invasion process. Sialidase-treated mouse erythrocytes were refractory to invasion by P. falciparum merozoites and failed to bind EBA-175. Influenza C virus esterase, which converts Neu5,9Ac2 to Neu5Ac, increased both invasion efficiency and EBA-175 binding to mouse erythrocytes. Thus, the parasite and EBA-175 discriminate between Neu5Ac and Neu5,9Ac2, that is, the C-9 acetyl group interferes with EBA-175 binding and invasion by P. falciparum merozoites. This indicates that sialic acid is part of a receptor for invasion.
Mol Biochem Parasitol 1992 Mar
PMID:Binding of Plasmodium falciparum 175-kilodalton erythrocyte binding antigen and invasion of murine erythrocytes requires N-acetylneuraminic acid but not its O-acetylated form. 156 37

The Salmonella typhimurium LT2 sialidase (neuraminidase, EC 3.2.1.18) structural gene, nanH, has been cloned and sialidase overproduced from multicopy plasmids in Escherichia coli. Sialidase expression was regulated positively by cAMP. In contrast, certain Tn1000 insertions located upstream of nanH coding sequences reduced sialidase activity. A nanH chromosomal insertion mutation constructed by marker exchange demonstrated a single sialidase gene copy in S. typhimurium LT2. The complete nucleotide sequence of nanH, encoding a 41,300 dalton polypeptide, was determined and the derived primary structure was similar to sialidases from Clostridium perfringens, Clostridium sordellii, Bacteroides fragilis, and Trypanosoma cruzi. Comparative sequence analysis, including codon usage and secondary structure predictions, indicated that the S. typhimurium and clostridial sialidases are homologous, strongly suggestive of an interspecies gene transfer event. At least two primary sequence motifs of the bacterial enzymes were detected in influenza A virus sialidases. The predicted secondary structure of the bacterial enzymes was strikingly similar to viral sialidase. From the population distribution of nanH detected within a collection of salmonellae, it was apparent that S. typhimurium obtained its nanH copy most recently from Salmonella arizonae. S. typhimurium LT2 is thus a genetic mosaic that differs from other strains of even the same serotype by nanH plus potentially additional characters linked to nanH. These results have relevance to the evolution and function of sialidases in pathogenic microbes, and to the origin of the sialic acids.
Mol Microbiol 1992 Apr
PMID:Cloning, sequencing and distribution of the Salmonella typhimurium LT2 sialidase gene, nanH, provides evidence for interspecies gene transfer. 160 67

An oligonucleotide mixture corresponding to the codons for conserved and repeated amino acid sequences of bacterial sialidases (Roggentin et al. 1989) was used to clone a 4.3 kb PstI restriction fragment of Clostridium septicum DNA in Escherichia coli. The complete nucleotide sequence of the sialidase gene was determined from this fragment. The derived amino acid sequence corresponds to a protein of 110,000 Da. The ribosomal binding site and promoter-like consensus sequences were identified upstream from the putative ATG initiation codon. The molecular and immunological properties of the sialidase expressed by E. coli are similar to those of the sialidase as isolated from C. septicum. The newly synthesized protein is assumed to include a leader peptide of 26 amino acids. On sequence alignment, the sialidases from C. septicum, C. sordellii and C. perfringens show significant homologies. As in other bacterial sialidases, conserved amino acid sequences occur at four positions in the protein. Aside from the consensus sequences, only poor homology to other bacterial and viral sialidases was found. The consensus sequence could be identified even in other, non-sialidase proteins, indicating a common function or the evolutionary relatedness of these proteins.
Mol Gen Genet 1991 Apr
PMID:The sialidase gene from Clostridium septicum: cloning, sequencing, expression in Escherichia coli and identification of conserved sequences in sialidases and other proteins. 203 13

We had previously shown that the human colon produces at least two immunochemically distinct mucins, one neutral and the other a sialomucin [Gold et al. J. biol. Chem. 256, 6354-6358 (1981)]. In addition, the sialomucin was shown to contain an immunodeterminant restricted to colonic epithelium and may thus prove useful as a tissue-specific marker. In the current study we have shown that a specific linkage of sialic acid to the oligosaccharide backbone has a major role in the organ-specific immunodeterminant structure. Treatment of intact colonic mucin with sialidase (Cl. perfringens) cleaved 20-80% of the sialic acid as measured colorimetrically. Immunoreactivity was decreased by 0-42% with respect to the untreated material. Saponification (0.1 N KOH, 20 min at room temp) caused an approximate 90% decrease in immunoreactivity for each mucin. Subsequent to saponification, neuraminidase cleaved most of the sialic acid from the mucins. The majority of sialic acid was observed to be O-acetylated, thus making it sialidase-insensitive. Gas chromatography-mass spectrometric analyses of the trimethylsilyl sialic acid derivatives indicated the presence of NeuNAc; NeuNAc, 9-OAc; and NeuNAc, 7,9 diOAc as the major sialyl derivatives. The radioimmunoassay data appeared to indicate that O-acetylated sialic acid was necessary for immunoreactivity. It should be noted that jejunal mucin and bovine submaxillary mucin also contain O-acetylated sialic acid, but did not inhibit in our radioimmunoassay. This may have been due to differences in the O-acetylation pattern or the linkage of sialic acid to the core carbohydrate. Analyses of the partially methylated alditol acetate derivatives by gas chromatography-mass spectrometry of the untreated, as well as the saponified and neuraminidase treated, mucins revealed that sialic acid was attached to the carbohydrate core either to galactose, N-acetylglucosamine, and/or N-acetylgalactosamine. Linear regression analyses comparing immunoreactivity with specific epitope concns, in conjunction with RIA analyses of known structures, suggested that the organ-specific immunodeterminant was (or was dependent upon the presence of) the structure GlcNAc (1,3)[O-acetylated Neu5Ac(2,6)] GalNAc.
Mol Immunol 1989 Aug
PMID:Studies on the structure of the organ-specific determinant of human colonic mucin. 247 76

Baso-lateral membranes were isolated from the canine and porcine kidney cortex by several different methods currently in use. Sidedness of the isolated membrane vesicles was determined by procedures using 1. ouabain-sensitive (Na+K+)ATPase assays in the presence and in the absence of sodium dodecylsulfate or digitoxigenin plus monensin, 2. (Na+, K+, Mg2+)ATPase assays with valinomycin, 3. sialidase accessibility, and 4. binding of hydrophilic and lipophilic cardiac glycosides. The (Na+K+)ATPase activity in the membrane preparation was increased 10-fold of that found in the crude homogenate. Isolated membrane vesicles, prepared by different techniques, were all found to be overwhelmingly of right-side-out orientation;namely, right-side-out = 51-68%, inside-out = 4-13%, and unsealed vesicles = 26-42%. Results of sidedness determinations by different methods showed a good agreement. Thus, predominantly right-side-out oriented vesicles are formed during conventional isolation procedures for membranes of the kidney cortex.
Mol Cell Biochem 1987 Nov
PMID:Orientation of vesicles isolated from baso-lateral membranes of renal cortex. 284 58

The carbohydrate antigen 19-9 (CA19-9) is considered to be of great importance in the diagnosis, differential diagnosis and follow-up of human pancreatic carcinoma. CA19-9 antigen has been isolated and characterized as the oligosaccharide sialylazed lacto-N-fucopentaose II and a monoclonal antibody against CA19-9 is commercially available. In this immunochemical study we have examined the localisation and distribution of monoclonal anti-CA19-9 in pancreatic tissue obtained from 20 patients with a normal pancreas (lacking pancreatic tumour or evidence of inflammation), from 50 patients with chronic pancreatitis and from 50 patients with pancreatic carcinomas of various types. In the normal pancreas (free from tumour or inflammation) we found anti-CA19-9 to be localized in the branches of the pancreatic ducts with discontinuities predominantly at the apical surfaces of the lining epithelium. In chronic pancreatitis a continuous positive reaction was found in the small, medium and large ramifications of the pancreatic ducts. In ductal epithelium exhibiting mucoid transformation, a mosaic-like, discontinuous positive reaction was found, whereas in epithelium showing pseudopapillary and papillary hyperplasia a uniform positive reaction was obtained. Multilayered epithelium ("squamous metaplasia") was negative. The fluid content of any cysts present and the tubular accumulations found in chronic pancreatitis showed a positive reaction. The reaction in chronic pancreatitis differed from that in normal pancreas in its distribution but not in its intensity. All carcinomas of the exocrine pancreas showed intensely positive reaction in a very varied distribution whereas the anaplastic carcinomas gave a negative reaction. Whilst in chronic pancreatitis the binding of anti-CA19-9 was unimpressive and strictly localized, in exocrine pancreatic carcinomas binding was and strictly localized, in exocrine pancreatic carcinomas binding was very marked and diffuse in distribution. From this we conclude that malignant cells display a greater number of CA19-9 epitopes than cells in chronic pancreatitis. The difference can only be regarded as quantitative, since the immunohistochemical reaction does not allow qualitative discrimination between chronic pancreatitis and pancreatic carcinoma; CA19-9 should not be therefore termed a "tumour marker". The glycoprotein nature of CA19-9 was confirmed by sialidase and chemical desialylation.
Virchows Arch B Cell Pathol Incl Mol Pathol 1986
PMID:The distribution and localization of the monoclonal antibody-defined antigen 19-9 (CA19-9) in chronic pancreatitis and pancreatic carcinoma. An immunohistochemical study. 287 26

During the course of their recirculation through the body, blood-borne lymphocytes specifically adhere to high endothelial venules (HEV) within secondary lymphoid organs such as peripheral lymph nodes (PN) and gut-associated Peyer's patches (PP). This adherence event, which initiates the extravasation of the lymphocyte, is highly specific in terms of the class of lymphocyte and the anatomic location of the HEV. We review evidence that the lymphocyte adhesive molecule ('homing receptor') involved in attachment to PN HEV is a carbohydrate-binding receptor (lectin-like) with specificity for mannose-6-phosphate (M6P)-like ligands. We describe the use of a novel cytochemical probe for the detection and characterization of cell surface carbohydrate-binding receptors. Using a M6P-based probe, we show that the carbohydrate-binding receptor on lymphocytes is closely-related or identical to the MEL-14 antigen, a putative homing receptor identified by a monoclonal antibody. Evidence is presented that the lymphocyte attachment sites on both PN and PP HEV are inactivated by mild periodate oxidation and hence are probably carbohydrate in nature. Yet, the sites are biochemically distinguishable in that one class (PN) requires sialidase-sensitive structures whereas the other (PP) does not. We raise the possibility that diversity in the carbohydrate-based recognition determinants on HEV may underlie the adhesive specificities in this system.
Mol Cell Biochem
PMID:Lymphocyte attachment to high endothelial venules during recirculation: a possible role for carbohydrates as recognition determinants. 302 59


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