Showing papers on "Sialic acid published in 2001"
TL;DR: The results suggest that AAV4 and AAV5 require different sialic acid-containing glycoproteins for binding and transduction of target cells and they further explain the different tropism of AAV 4 and A AV5.
Abstract: Adeno-associated virus serotype 4 (AAV4) and AAV5 have different tropisms compared to AAV2 and to each other. We recently reported that α2-3 sialic acid is required for AAV5 binding and transduction. In this study, we characterized AAV4 binding and transduction and found it also binds sialic acid, but the specificity is significantly different from AAV5. AAV4 can hemagglutinate red blood cells from several species, whereas AAV5 hemagglutinates only rhesus monkey red blood cells. Treatment of red blood cells with trypsin inhibited hemagglutination for both AAV4 and AAV5, suggesting that the agglutinin is a protein. Treatment of Cos and red blood cells with neuraminidases also indicated that AAV4 bound α2-3 sialic acid. However, resialylation experiments with neuraminidase-treated red blood cells demonstrated that AAV4 binding required α2–3 O-linked sialic acid, whereas AAV5 required N-linked sialic acid. Similarly, resialylation of sialic acid-deficient CHO cells supported this same conclusion. The difference in linkage specificity for AAV4 and AAV5 was confirmed by binding and transduction experiments with cells incubated with either N-linked or O-linked inhibitors of glycosylation. Furthermore, AAV4 transduction was only blocked with soluble α2-3 sialic acid, whereas AAV5 could be blocked with either α2–3 or α2-6 sialic acid. These results suggest that AAV4 and AAV5 require different sialic acid-containing glycoproteins for binding and transduction of target cells and they further explain the different tropism of AAV4 and AAV5.
436 citations
TL;DR: The crystal structures of Newcastle disease virus HN alone and in complex with either an inhibitor or with the β-anomer of sialic acid reveal a typical neuraminidase active site within a β-propeller fold.
Abstract: Paramyxoviruses are the main cause of respiratory disease in children. One of two viral surface glycoproteins, the hemagglutinin-neuraminidase (HN), has several functions in addition to being the major surface antigen that induces neutralizing antibodies. Here we present the crystal structures of Newcastle disease virus HN alone and in complex with either an inhibitor or with the β-anomer of sialic acid. The inhibitor complex reveals a typical neuraminidase active site within a β-propeller fold. Comparison of the structures of the two complexes reveal differences in the active site, suggesting that the catalytic site is activated by a conformational switch. This site may provide both sialic acid binding and hydrolysis functions since there is no evidence for a second sialic acid binding site in HN. Evidence for a single site with dual functions is examined and supported by mutagenesis studies. The structure provides the basis for the structure-based design of inhibitors for a range of paramyxovirus-induced diseases.
386 citations
TL;DR: It is found that in contrast to AAV2, the apical membrane of airway epithelia contains abundant high affinity receptors for AAV5, and observations suggest that 2,3-linked sialic acid is either a receptor for A AV5 or it is a necessary component of a receptor complex.
362 citations
TL;DR: Evidence that the human condition can explain differences in susceptibility or resistance to certain microbial pathogens and a markedly reduced expression of hydroxylase in the brains of other mammals raises the possibility that thehuman‐specific mutation of this enzyme could have played a role in human brain evolution.
Abstract: The surface of all mammalian cells is covered with a dense and complex array of sugar chains, which are frequently terminated by members of a family of molecules called sialic acids. One particular sialic acid called N-glycolylneuraminic acid (Neu5Gc) is widely expressed on most mammalian tissues, but is not easily detectable on human cells. In fact, it provokes an immune response in adult humans. The human deficiency of Neu5Gc is explained by an inactivating mutation in the gene encoding CMP-N-acetylneuraminic acid hydroxylase, the rate-limiting enzyme in generating Neu5Gc in cells of other mammals. This deficiency also results in an excess of the precursor sialic acid N-acetylneuraminic acid (Neu5Ac) in humans. This mutation appears universal to modern humans, occurred sometime after our last common ancestor with the great apes, and happens to be one of the first known human-great ape genetic differences with an obvious biochemical readout. While the original selection mechanisms and major biological consequences of this human-specific mutation remain uncertain, several interesting clues are currently being pursued. First, there is evidence that the human condition can explain differences in susceptibility or resistance to certain microbial pathogens. Second, the functions of some endogenous receptors for sialic acids in the immune system may be altered by this difference. Third, despite the lack of any obvious alternate pathway for synthesis, Neu5Gc has been reported in human tumors and possibly in human fetal tissues, and traces have even been detected in normal human tissues. One possible explanation is that this represents accumulation of Neu5Gc from dietary sources of animal origin. Finally, a markedly reduced expression of hydroxylase in the brains of other mammals raises the possibility that the human-specific mutation of this enzyme could have played a role in human brain evolution.
290 citations
TL;DR: The expression pattern of human sialoadhesin was found to be similar to that of the mouse receptor, being absent from monocytes and other peripheral blood leukocytes, but expressed strongly by tissue macrophages in the spleen, lymph node, bone marrow, liver, colon, and lungs.
289 citations
TL;DR: A critical review of the evidence for Neu5Gc in human tumours is provided, paying particular attention to the analytical methods employed, and alternative metabolic pathways for the biosynthesis of glycoconjugate-bound Neu 5Gc are considered.
271 citations
TL;DR: The side chain of sialic acid offers new tools to study its biological relevance and to exploit it as a tag for therapeutic and diagnostic applications as mentioned in this paper, including its crucial role for the interaction of different viruses with their sialylated host cell receptors.
Abstract: N-Acetylneuraminic acid is the most prominent sialic acid in eukaryotes. The structural diversity of sialic acid is exploited by viruses, bacteria, and toxins and by the sialoglycoproteins and sialoglycolipids involved in cell-cell recognition in their highly specific recognition and binding to cellular receptors. The physiological precursor of all sialic acids is N-acetyl D-mannosamine (ManNAc). By recent findings it could be shown that synthetic N-acyl-modified D-mannosamines can be taken up by cells and efficiently metabolized to the respective N-acyl-modified neuraminic acids in vitro and in vivo. Successfully employed D-mannosamines with modified N-acyl side chains include N-propanoyl- (ManNProp), N-butanoyl- (ManNBut)-, N-pentanoyl- (ManNPent), N-hexanoyl- (ManNHex), N-crotonoyl- (ManNCrot), N-levulinoyl- (ManNLev), N-glycolyl- (ManNGc), and N-azidoacetyl D-mannosamine (ManNAc-azido). All of these compounds are metabolized by the promiscuous sialic acid biosynthetic pathway and are incorporated into cell surface sialoglycoconjugates replacing in a cell type-specific manner 10-85% of normal sialic acids. Application of these compounds to different biological systems has revealed important and unexpected functions of the N-acyl side chain of sialic acids, including its crucial role for the interaction of different viruses with their sialylated host cell receptors. Also, treatment with ManNProp, which contains only one additional methylene group compared to the physiological precursor ManNAc, induced proliferation of astrocytes, microglia, and peripheral T-lymphocytes. Unique, chemically reactive ketone and azido groups can be introduced biosynthetically into cell surface sialoglycans using N-acyl-modified sialic acid precursors, a process offering a variety of applications including the generation of artificial cellular receptors for viral gene delivery. This group of novel sialic acid precursors enabled studies on sialic acid modifications on the surface of living cells and has improved our understanding of carbohydrate receptors in their native environment. The biochemical engineering of the side chain of sialic acid offers new tools to study its biological relevance and to exploit it as a tag for therapeutic and diagnostic applications.
271 citations
TL;DR: The presence of two conserved immunoreceptor tyrosine-based inhibition motif (ITIM)-like motifs in their cytoplasmic tails suggests that, like CD22, CD33-related Siglecs are involved in regulating cellular activation within the immune system.
Abstract: Siglecs1 (sialic acid binding Ig-like lectins) are I-type (Ig-type) lectins2 characterized by an N-terminal V-set Ig domain that mediates sialic acid binding,3 followed by varying numbers of C2-set Ig domains (Fig. 1). The initial discovery of this lectin family came about through independent studies on sialoadhesin (Siglec-1/CD169), a macrophage lectin-like adhesion molecule,4 and CD22 (Siglec-2), a B-cell restricted member of the Ig superfamily (IgSF)5 that plays an important role in regulating B-cell activation. Both molecules were found to mediate cell–cell interactions in vitro via recognition of sialylated glycoconjugates.6–10 The cloning of sialoadhesin11 revealed striking sequence similarities to CD22 and led to the demonstration that two other related IgSF proteins, myelin-associated glycoprotein (MAG/Siglec-4) and CD33 (Siglec-3), which were not previously known to bind sialic acids, were also members of the Siglec family (Table 1).12,13
Figure 1
Structural features of Siglecs. (a) Siglecs are type I membrane proteins with an extracellular region containing a sialic acid binding V-set Ig-like domain at the N-terminus and 1–16 C2-set Ig-like domains. The cytoplasmic tails of all Siglecs ...
Table 1
Properties of Siglecs
Six additional human Siglecs (Siglecs 5–10) have been identified and characterized over the last 3 years. These previously unknown molecules share a high degree of sequence similarity with CD33 in their extracellular and intracellular regions, and are hence collectively referred to as ‘CD33-related Siglecs’. A striking feature of the CD33-related Siglecs is their differential expression pattern amongst the cell lineages of the haemopoietic system (see Table 1). This, together with the presence of two conserved immunoreceptor tyrosine-based inhibition motif (ITIM)-like motifs in their cytoplasmic tails, suggests that, like CD22, CD33-related Siglecs are involved in regulating cellular activation within the immune system. Here we discuss how sialic acid recognition by Siglecs might contribute to the regulation of immune functions.
248 citations
01 Jan 2001
TL;DR: Application of these compounds to different biological systems has revealed important and unexpected functions of the N-acyl side chain of sialic acids, including its crucial role for the interaction of different viruses with their sialylated host cell receptors.
Abstract: viral gene delivery. This group of novel sialic acid precursors enabled studies on sialic acid modifications on the surface of living cells and has improved our understanding of carbohydrate receptors in their native environment. The biochemical engineering of the side chain of sialic acid offers new tools to study its biological relevance and to exploit it as a tag for therapeutic and diagnostic applications.
233 citations
TL;DR: Human milk, including milk from mothers of preterm infants, is a rich source of oligosaccharide-bound sialic acid, which contrasts with the relatively small amounts found in infant formulas.
230 citations
TL;DR: It is demonstrated unequivocally that gangliosides play an essential role in the proper functioning of the central nervous system (CNS) and that GD3S−/− mice, even with an absence of b-series ganglioides, appear to undergo normal development and have a normal life span.
TL;DR: In this paper, the authors examined the feasibility of implementing metabolic control strategies aimed at reducing the occurrence of murine-specific glycan motifs on NS0-derived recombinant proteins, such as Galalpha1,3Galbeta1,4GlcNAc.
Abstract: Chinese hamster ovary and murine myeloma NS0 cells are currently favored host cell types for the production of therapeutic recombinant proteins. In this study, we compared N-glycan processing in GS-NS0 and GS-CHO cells producing the same model recombinant glycoprotein, tissue inhibitor of metalloproteinases 1. By manipulation of intracellular nucleotide-sugar content, we examined the feasibility of implementing metabolic control strategies aimed at reducing the occurrence of murine-specific glycan motifs on NS0-derived recombinant proteins, such as Galalpha1,3Galbeta1,4GlcNAc. Although both CHO and NS0-derived oligosaccharides were predominantly of the standard complex type with variable sialylation, 30% of N-glycan antennae associated with NS0-derived TIMP-1 terminated in alpha1,3-linked galactose residues. Furthermore, NS0 cells conferred a greater proportion of terminal N-glycolylneuraminic (sialic) acid residues as compared with the N-acetylneuraminic acid variant. Inclusion of the nucleotide-sugar precursors, glucosamine (10 mM, plus 2 mM uridine) and N-acetylmannosamine (20 mM), in culture media were shown to significantly increase the intracellular pools of UDP-N-acetylhexosamine and CMP-sialic acid, respectively, in both NS0 and CHO cells. The elevated UDP-N-acetylhexosamine content induced by the glucosamine/uridine treatment was associated with an increase in the antennarity of N-glycans associated with TIMP-1 produced in CHO cells but not N-glycans associated with TIMP-1 from NS0 cells. In addition, elevated UDP-N-acetylhexosamine content was associated with a slight decrease in sialylation in both cell lines. The elevated CMP-sialic acid content induced by N-acetylmannosamine had no effect on the overall level of sialylation of TIMP-1 produced by both CHO and NS0 cells, although the ratio of N-glycolylneuraminic acid:N-acetylneuraminic acid associated with NS0-derived TIMP-1 changed from 1:1 to 1:2. These data suggest that manipulation of nucleotide-sugar metabolism can promote changes in N-glycan processing that are either conserved between NS0 and CHO cells or specific to either NS0 cells or CHO cells.
TL;DR: Results indicate that reovirus binding to sialic acid enhances virus infection through adhesion of virus to the cell surface where access to a proteinaceous receptor is thermodynamically favored.
TL;DR: Even when C2GnT1 is expressed, the O-glycans added to MUC1 become core 1-dominated structures, provided expression of ST3Gal-I is increased as it is in breast cancer, which indicates a switch to core 2 structures.
TL;DR: It appears that if insect cells have the genetic potential to perform sialylation of glycoproteins, this is a highly specialized function that probably occurs rarely and will require metabolic engineering efforts to extend the native protein glycosylation pathways of insect cells.
Abstract: Our growing comprehension of the biological roles of glycan moieties has created a clear need for expression systems that can produce mammalian-type glycoproteins. In turn, this has intensified interest in understanding the protein glycosylation pathways of the heterologous hosts that are commonly used for recombinant glycoprotein expression. Among these, insect cells are the most widely used and, particularly in their role as hosts for baculovirus expression vectors, provide a powerful tool for biotechnology. Various studies of the glycosylation patterns of endogenous and recombinant glycoproteins produced by insect cells have revealed a large variety of O- and N-linked glycan structures and have established that the major processed O- and N-glycan species found on these glycoproteins are (Gal beta1,3)GalNAc-O-Ser/Thr and Man3(Fuc)GlcNAc2-N-Asn, respectively. However, the ability or inability of insect cells to synthesize and compartmentalize sialic acids and to produce sialylated glycans remains controversial. This is an important issue because terminal sialic acid residues play diverse biological roles in many glycoconjugates. While most work indicates that insect cell-derived glycoproteins are not sialylated, some well-controlled studies suggest that sialylation can occur. In evaluating this work, it is important to recognize that oligosaccharide structural determination is tedious work, due to the infinite diversity of this class of compounds. Furthermore, there is no universal method of glycan analysis; rather, various strategies and techniques can be used, which provide glycobiologists with relatively more or less precise and reliable results. Therefore, it is important to consider the methodology used to assess glycan structures when evaluating these studies. The purpose of this review is to survey the studies that have contributed to our current view of glycoprotein sialylation in insect cell systems, according to the methods used. Possible reasons for the disagreement on this topic in the literature, which include the diverse origins of biological material and experimental artifacts, will be discussed. In the final analysis, it appears that if insect cells have the genetic potential to perform sialylation of glycoproteins, this is a highly specialized function that probably occurs rarely. Thus, the production of sialylated recombinant glycoproteins in the baculovirus-insect cell system will require metabolic engineering efforts to extend the native protein glycosylation pathways of insect cells.
TL;DR: In vitro galactosylation and sialylation of therapeutic glycoproteins with terminal GlcNAc and Gal residues can be achieved in a single step, and the results are similar to those for the stepwise reaction.
Abstract: Therapeutic glycoproteins produced in different host cells by recombinant DNA technology often contain terminal GlcNAc and Gal residues. Such glycoproteins clear rapidly from the serum as a consequence of binding to the mannose receptor and/or the asialoglycoprotein receptor in the liver. To increase the serum half-life of these glycoproteins, we carried out in vitro glycosylation experiments using TNFR-IgG, an immunoadhesin molecule, as a model therapeutic glycoprotein. TNFR-IgG is a disulfide-linked dimer of a polypeptide composed of the extracellular portion of the human type 1 (p55) tumor necrosis factor receptor (TNFR) fused to the hinge and Fc regions of the human IgG(1) heavy chain. This bivalent antibody-like molecule contains four N-glycosylation sites per polypeptide, three in the receptor portion and one in the Fc. The heterogeneous N-linked oligosaccharides of TNFR-IgG contain sialic acid (Sia), Gal, and GlcNAc as terminal sugar residues. To increase the level of terminal sialylation, we regalactosylated and/or resialylated TNFR-IgG using beta-1,4-galactosyltransferase (beta1,4GT) and/or alpha-2,3-sialyltransferase (alpha2,3ST). Treatment of TNFR-IgG with beta1,4GT and UDP-Gal, in the presence of MnCl(2), followed by MALDI-TOF-MS analysis of PNGase F-released N-glycans showed that the number of oligosaccharides with terminal GlcNAc residues was significantly decreased with a concomitant increase in the number of terminal Gal residues. Similar treatment of TNFR-IgG with alpha2,3ST and CMP-sialic acid (CMP-Sia), in the presence of MnCl(2), produced a molecule with an approximately 11% increase in the level of terminal sialylation but still contained oligosaccharides with terminal GlcNAc residues. When TNFR-IgG was treated with a combination of beta1,4GT and alpha2,3ST (either in a single step or in a stepwise fashion), the level of terminal sialylation was increased by approximately 20-23%. These results suggest that in vitro galactosylation and sialylation of therapeutic glycoproteins with terminal GlcNAc and Gal residues can be achieved in a single step, and the results are similar to those for the stepwise reaction. This type of in vitro glycosylation is applicable to other glycoproteins containing terminal GlcNAc and Gal residues and could prove to be useful in increasing the serum half-life of therapeutic glycoproteins.
TL;DR: Results suggest that EBA140 may be involved in merozoite invasion using a sialic acid‐dependent receptor on human erythrocytes, and a parasite line that lacks the eba140 gene, suggesting that this protein is not essential for in vitro invasion.
Abstract: Summary Invasion of the merozoite form of Plasmodium falciparum into human erythrocytes involves multiple receptor‐ligand interactions. The EBA175 protein of P. falciparum has been shown to be the ligand that binds to a sialic acid-dependent site on glycophorin A. We have identified a novel P. falciparum ligand, termed erythrocyte-binding antigen 140 (EBA140), that shares structural features and homology with EBA175. Subcellular localization of EBA140 suggests that it is located in the micronemes, the same localization as EBA175. EBA140 binds to a sialic acid-dependent receptor on the surface of human erythrocytes. Binding of EBA140 to this erythrocyte receptor is sensitive to neuraminidase and resistant to trypsin, proteinase K and pronase. The proteaseresistant properties of the erythrocyte receptor suggests that it is not glycophorin A or C. Additionally, analysis of mutant erythrocytes from humans has shown that EBA140 does not bind glycophorin B. Interestingly, we have identified a parasite line that lacks the eba140 gene, suggesting that this protein is not essential for in vitro invasion. These results suggest that EBA140 may be involved in merozoite invasion using a sialic acid-dependent receptor on human erythrocytes.
TL;DR: The resulting decrease in Gc concentration makes Gc usable as a prognostic indicator of survival of patients with significant tissue injury after trauma and among patients with hepatic failure, as well as among women of known Gc phenotype.
Abstract: Immunonephelometry has been reported (1) to be a suitable method for quantification of vitamin D-binding protein (also known as Gc globulin or Gc). We wished to develop such a method and examine the association between the mean serum concentration of Gc in women of known Gc phenotype and the phenotype of Gc.
Gc is a 52- to 58-kDa multifunctional plasma protein, synthesized mainly by hepatocytes. Polymorphisms in the Gc gene (codominant alleles) give rise to three major electrophoretic variants of Gc (Gc2, Gc1s, and Gc1f), which differ by amino acid substitutions as well as glycosylation (2)(3). The physiological significance related to the various phenotypes is yet to be discovered.
Gc is the major carrier protein of vitamin D and its metabolites in the circulation and is important for preservation of the vitamin (4)(5). Gc also transports components such as fatty acids and endotoxin (6)(7), and it is an important player in the actin scavenging system (8)(9). Gc binds actin released from cells upon injury, and the Gc-actin complexes are rapidly cleared from the circulation, thereby preventing the harmful effects of actin filaments in blood vessels. The resulting decrease in Gc concentration makes Gc usable as a prognostic indicator of survival of patients with significant tissue injury after trauma (10) and among patients with hepatic failure (11).
In addition to being a transporter and an actin scavenger protein, Gc may be of importance for bone formation and in the immune system. After in vitro removal of its galactose and sialic acid residues, Gc is converted to a very potent macrophage-activating factor, Gc-MAF (12). Administration of Gc-MAF to osteopetrotic rodents reversed their bone and immunological defects, probably by activating osteoclasts as well as macrophages (13). Finally, together with complement factors C5a and C5a …
TL;DR: CD43 is identified as a T cell counterreceptor for sialoadhesin and it is suggested that in addition to its anti-adhesive role CD43 may promote cell-cell interactions.
Abstract: Sialoadhesin (Siglec-1) is a macrophage-restricted sialic acid-binding receptor that mediates interactions with hemopoietic cells, including lymphocytes. In this study, we identify sialoadhesin counterreceptors on T lymphocytes. Several major glycoproteins (85, 130, 240 kDa) were precipitated by sialoadhesin-Fc fusion proteins from a murine T cell line (TK-1). Binding of sialoadhesin to these glycoproteins was sialic acid dependent and was abolished by mutation of a critical residue (R97A) of the sialic acid binding site in the membrane distal Ig-like domain of sialoadhesin. The 130- and 240-kDa sialoadhesin-binding glycoproteins were identified as the sialomucins CD43 and P-selectin glycoprotein ligand 1 (CD162), respectively. CD43 expressed in COS cells supported increased binding to immobilized sialoadhesin. Finally, sialoadhesin bound different glycoforms of CD43 expressed in Chinese hamster ovary cells, including unbranched (core 1) and branched (core 2) O:-linked glycans, that are normally found on CD43 in resting and activated T cells, respectively. These results identify CD43 as a T cell counterreceptor for sialoadhesin and suggest that in addition to its anti-adhesive role CD43 may promote cell-cell interactions.
TL;DR: A surface plasmon resonance (SPR) sensor using a molecularly imprinted polymer-coated sensor chip for the detection of sialic acid showed a selective response to ganglioside of which sIALic acid is located at the non-reducing end and gave a linear relationship from 0.1 to 1.0 mg of gangliosiside.
TL;DR: The binding of triple-layered particles of SA11 and NCDV to NeuGc-GM3, but not toNeuAc-GM 3, suggested that the sequence NeUGcα3Galβ is preferred to NeUAcα3 Galβ, which indicates neuraminidase-insensitive rotavirus strains recognize gangliosides with internal sialic acids, which are resistant to neuramidase treatment.
Abstract: The glycosphingolipid binding specificities of neuraminidase-sensitive (simian SA11 and bovine NCDV) and neuraminidase-insensitive (bovine UK) rotavirus strains were investigated using the thin-layer chromatogram binding assay. Both triple-layered and double-layered viral particles of SA11, NCDV, and UK bound to nonacid glycosphingolipids, including gangliotetraosylceramide (GA1; also called asialo-GM1) and gangliotriaosylceramide (GA2; also called asialo-GM2). Binding to gangliosides was observed with triple-layered particles but not with double-layered particles. The neuraminidase-sensitive and neuraminidase-insensitive rotavirus strains showed distinct ganglioside binding specificities. All three strains bound to sialylneolactotetraosylceramide and GM2 and GD1a gangliosides. However, NeuAc-GM3 and the GM1 ganglioside were recognized by rotavirus strain UK but not by strains SA11 and NCDV. Conversely, NeuGc-GM3 was bound by rotaviruses SA11 and NCDV but not by rotavirus UK. Thus, neuraminidase-sensitive strains bind to external sialic acid residues in gangliosides, while neuraminidase-insensitive strains recognize gangliosides with internal sialic acids, which are resistant to neuraminidase treatment. By testing a panel of gangliosides with triple-layered particles of SA11 and NCDV, the terminal sequence sialyl-galactose (NeuGc/NeuAcα3-Galβ) was identified as the minimal structural element required for the binding of these strains. The binding of triple-layered particles of SA11 and NCDV to NeuGc-GM3, but not to NeuAc-GM3, suggested that the sequence NeuGcα3Galβ is preferred to NeuAcα3Galβ. Further dissection of this binding epitope showed that the carboxyl group and glycerol side chain of sialic acid played an important role in the binding of such triple-layered particles.
TL;DR: The abilities of human parainfluenza virus types 1 and 3 to bind to different types of gangliosides are evaluated, indicating that the molecular species of glycoconjugate that hPIV-1 recognizes are more limited than those recognized by hPiv-3.
Abstract: Through their hemagglutinin-neuraminidase glycoprotein, parainfluenza viruses bind to sialic acid-containing glycoconjugates to initiate infection. Although the virus-receptor interaction is a key factor of infection, the exact nature of the receptors that human parainfluenza viruses recognize has not been determined. We evaluated the abilities of human parainfluenza virus types 1 (hPIV-1) and 3 (hPIV-3) to bind to different types of gangliosides. Both hPIV-1 and hPIV-3 preferentially bound to neolacto-series gangliosides containing a terminal N-acetylneuraminic acid (NeuAc) linked to N-acetyllactosamine (Galβ1-4GlcNAc) by the α2-3 linkage (NeuAcα2-3Galβ1-4GlcNAc). Unlike hPIV-1, hPIV-3 bound to gangliosides with a terminal NeuAc linked to Galβ1-4GlcNAc through an α2-6 linkage (NeuAcα2-6Galβ1-4GlcNAc) or to gangliosides with a different sialic acid, N-glycolylneuraminic acid (NeuGc), linked to Galβ1-4GlcNAc (NeuGcα2-3Galβ1-4GlcNAc). These results indicate that the molecular species of glycoconjugate that hPIV-1 recognizes are more limited than those recognized by hPIV-3. Further analysis using purified gangliosides revealed that the oligosaccharide core structure is also an important element for binding. Gangliosides that contain branched N-acetyllactosaminoglycans in their core structure showed higher avidity than those without them. Agglutination of human, cow, and guinea pig erythrocytes but not equine erythrocytes by hPIV-1 and hPIV-3 correlated well with the presence or the absence of sialic acid-linked branched N-acetyllactosaminoglycans on the cell surface. Finally, NeuAcα2-3I, which bound to both viruses, inhibited virus infection of Lewis lung carcinoma-monkey kidney cells in a dose-dependent manner. We conclude that hPIV-1 and hPIV-3 preferentially recognize oligosaccharides containing branched N-acetyllactosaminoglycans with terminal NeuAcα2-3Gal as receptors and that hPIV-3 also recognizes NeuAcα2-6Gal- or NeuGcα2-3Gal-containing receptors. These findings provide important information that can be used to develop inhibitors that prevent human parainfluenza virus infection.
TL;DR: It is found that the human deficiency of Neu5Gc can be explained by an inactivating mutation in the gene encoding CMP-N-acetylneuraminic acid hydroxylase, and thus Neu 5Gc is not an oncofetal antigen in the classical sense, and other explanations must be found for the observed expression pattern.
TL;DR: This report describes the investigation into the extent of tolerance of the pathway toward additional structural alterations of the N-acyl substituent of ManNAc, and the results suggested that cellular metabolism is limited by the phosphorylation ofThe N-ACYlmannosamines by Man NAc 6-kinase in the first step of the pathways.
Abstract: Unnatural analogues of sialic acid can be delivered to mammalian cell surfaces through the metabolic transformation of unnatural N-acetylmannosamine (ManNAc) derivatives. In previous studies, mannosamine analogues bearing simple N-acyl groups up to five carbon atoms in length were recognized as substrates by the biosynthetic machinery and transformed into cell surface sialoglycoconjugates [Keppler, O. T., et al. (2001) Glycobiology 11, 11R-18R]. Such structural alterations to cell surface glycans can be used to probe carbohydrate-dependent phenomena. This report describes our investigation into the extent of tolerance of the pathway toward additional structural alterations of the N-acyl substituent of ManNAc. A panel of analogues with ketone-containing N-acyl groups that varied in the length or steric bulk was chemically synthesized and tested for metabolic conversion to cell surface glycans. We found that extension of the N-acyl chain to six, seven, or eight carbon atoms dramatically reduced utilization by the biosynthetic machinery. Likewise, branching from the linear chain reduced metabolic conversion. Quantitation of metabolic intermediates suggested that cellular metabolism is limited by the phosphorylation of the N-acylmannosamines by ManNAc 6-kinase in the first step of the pathway. This was confirmed by enzymatic assay of the partially purified enzyme with unnatural substrates. Identification of ManNAc 6-kinase as a bottleneck for unnatural sialic acid biosynthesis provides a target for expanding the metabolic promiscuity of mammalian cells.
TL;DR: A recombinant IgG3 antibody with Phe-243 replaced by Ala (FA243) having both alpha2,6 andalpha2,3-sialylation restored recognition to wild-type IgG 3 levels for human FcgammaRI, FcGammaRII, and target cell lysis by complement.
TL;DR: It is demonstrated that reovirus binding to cell surface sialic acid is a critical requirement for the efficient induction of apoptosis and suggested that virus receptor utilization plays an important role in regulating cell death.
Abstract: Reovirus induces apoptosis in cultured cells and in vivo. Genetic studies indicate that the efficiency with which reovirus strains induce apoptosis is determined by the viral S1 gene, which encodes attachment protein ς1. However, the biochemical properties of ς1 that influence apoptosis induction are unknown. To determine whether the capacity of ς1 to bind cell surface sialic acid determines the magnitude of the apoptotic response, we used isogenic reovirus mutants that differ in the capacity to engage sialic acid. We found that T3SA+, a virus capable of binding sialic acid, induces high levels of apoptosis in both HeLa cells and L cells. In contrast, non-sialic-acid-binding strain T3SA− induces little or no apoptosis in these cell types. Differences in the capacity of T3SA− and T3SA+ to induce apoptosis are not due to differences in viral protein synthesis or production of viral progeny. Removal of cell surface sialic acid with neuraminidase abolishes the capacity of T3SA+ to induce apoptosis. Similarly, incubation of T3SA+ with sialyllactose, a trisaccharide comprised of lactose and sialic acid, blocks apoptosis. These findings demonstrate that reovirus binding to cell surface sialic acid is a critical requirement for the efficient induction of apoptosis and suggest that virus receptor utilization plays an important role in regulating cell death.
TL;DR: This review summarizes the current research on human exo -α-sialidase (sIALidase, neuraminidase).
Abstract: This review summarizes the current research on human exo-alpha-sialidase (sialidase, neuraminidase). Where appropriate, the properties of viral, bacterial, and human sialidases have been compared. Sialic acids are implicated in diverse physiological processes. Sialidases, as enzymes acting upon sialic acids, assume importance as well. Sialidases hydrolyze the terminal, non-reducing, sialic acid linkage in glycoproteins, glycolipids, gangliosides, polysaccharides, and synthetic molecules. Therefore, a variety of assays are available to measure sialidase activity. Human sialidase is present in several organs and cells. Its cellular distribution could be cytosolic, lysosomal, or in the membrane. Human sialidase occurs in a high molecular-mass complex with several other proteins, including cathepsin A and beta-galactosidase. Multi-protein complexation is important for the in vivo integrity and catalytic activity of the sialidase. However, multi-protein complexation, the occurrence of isoenzymes, diverse subcellular localization, thermal instability, and membrane association have all contributed to difficulties in purifying and characterizing human sialidases. Human sialidase isoenzymes have recently been cloned and sequenced. Even though crystal structures for the human sialidases are not available, the highly conserved regions of the sialidase from various organisms have facilitated molecular modeling of the human enzyme and raise interesting evolutionary questions. While the molecular mechanisms vary, genetic defects leading to human sialidase deficiency are closely associated with at least two well-known human diseases, namely sialidosis and galactosialidosis. No therapy is currently available for either disease. A thorough investigation of human sialidases is therefore crucial to human health.
TL;DR: 5-Azido sialyl donors with O-acetyl protecting groups are useful α-selective glycosylation reagents, especially for primary hydroxyl groups as acceptors, and it was also possible to synthesize NeuAcα(2→9)NeuAc as a thioglycoside donor for use in subsequent gly cosylations.
Abstract: 5-Azido sialyl donors with O-acetyl protecting groups are useful α-selective glycosylation reagents, especially for primary hydroxyl groups as acceptors. This is shown with a variety of reactions using 1 as a sialyl donor. It was also possible to synthesize NeuAcα(2→9)NeuAc as a thioglycoside donor for use in subsequent glycosylations.
TL;DR: The effects of cholesterol and sialic acid depletion on Aβ-induced GTPase activity in cells, a step implicated in the mechanism of Aβ toxicity, and A β-induced cell toxicity are investigated and control of cellular cholesterol and/or ganglioside content may prove useful in the prevention or treatment of Alzheimer's disease.
TL;DR: Among all the peptidic fragments obtained by tryptic digestion of bLF and characterised by advanced mass spectrometric methodologies, a large fragment and a small peptide were able to inhibit rotavirus even if at lower extent than undigested bLf.