Expression of a functional Drosophila melanogasterN-acetylneuraminic acid (Neu5Ac) phosphate synthase gene: evidence for endogenous sialic acid biosynthetic ability in insects

The first step in influenza virus infection is the binding of hemagglutinin to sialic acid-containing glycans present on the cell surface. Over 50 different sialic acid modifications are known, of which N-acetylneuraminic acid (Neu5Ac) and N-glycolylneuraminic acid (Neu5Gc) are the two main species. Animal models with α2,6 linked Neu5Ac in the upper respiratory tract, similar to humans, are preferred to enable and mimic infection with unadapted human influenza A viruses. Animal models that are currently most often used to study human influenza are mice and ferrets. Additionally, guinea pigs, cotton rats, Syrian hamsters, tree shrews, domestic swine, and non-human primates (macaques and marmosets) are discussed. The presence of NeuGc and the distribution of sialic acid linkages in the most commonly used models is summarized and experimentally determined. We also evaluated the role of Neu5Gc in infection using Neu5Gc binding viruses and cytidine monophosphate-N-acetylneuraminic acid hydroxylase (CMAH)-/- knockout mice, which lack Neu5Gc and concluded that Neu5Gc is unlikely to be a decoy receptor. This article provides a base for choosing an appropriate animal model. Although mice are one of the most favored models, they are hardly naturally susceptible to infection with human influenza viruses, possibly because they express mainly α2,3 linked sialic acids with both Neu5Ac and Neu5Gc modifications. We suggest using ferrets, which resemble humans closely in the sialic acid content, both in the linkages and the lack of Neu5Gc, lung organization, susceptibility, and disease pathogenesis.

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Interaction of Mycoplasma gallisepticum with sialyl glycoproteins

Infection and Immunity ◽

10.1128/iai.30.2.353-361.1980 ◽

1980 ◽

Vol 30 (2) ◽

pp. 353-361

Author(s):

L R Glasgow ◽

R L Hill

Keyword(s):

Sialic Acid ◽

Mycoplasma Gallisepticum ◽

Binding Assays ◽

Molecular Weights ◽

Acetylneuraminic Acid ◽

Direct Binding ◽

Membrane Bound ◽

Strict Specificity ◽

Human Glycophorin ◽

Acid Structure

The binding of several glycoproteins to freshly grown and harvested cells of Mycoplasma gallisepticum was examined. Only human glycophorin, the major sialoglycoprotein of the erythrocyte membrane, bound tightly as judged by direct binding assays with 125I-labeled glycoproteins. Neuraminidase-treated glycophorin did not bind, suggesting that binding is mediated through sialic acid groups. Although other sialoglycoproteins did not appear to bind M. gallisepticum by direct binding assays, some inhibited the binding of glycophorin. The best inhibitors had a mucin-like structure, with high molecular weights and high sialic acid contents. N-acetylneuraminic acid appeared to be the favored sialic acid structure for binding, but there was no strict specificity for its anomeric linkage. Neuraminidase activity could not be detected on the surface of M. gallisepticum, suggesting that this enzyme is not involved in the mechanism of adherence of sialoglycoproteins. Binding of sialoglycoproteins was time dependent, however, and markedly diminished with increasing ionic strength, but was largely unaffected between pH 4 and 9.

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Porcine Sugar Nucleotide: Glycoprotein Glycosyltransferases. I. Blood Serum and Liver Sialyltransferase

Canadian Journal of Biochemistry ◽

10.1139/o71-117 ◽

1971 ◽

Vol 49 (7) ◽

pp. 829-837 ◽

Cited By ~ 64

Author(s):

Roger L. Hudgin ◽

Harry Schachter

Keyword(s):

Sialic Acid ◽

High Speed ◽

Liver Homogenate ◽

Acetone Powder ◽

Acid Glycoprotein ◽

Acetylneuraminic Acid ◽

Pork Liver ◽

Terminal Galactose ◽

And Function ◽

Powder Extract

The properties of CMP-N acetylneuraminic acid: glycoprotein sialyltransferase have been studied in pork serum, a crude pork liver homogenate, and a soluble acetone powder extract prepared from pork liver. Whereas the crude liver homogenate enzyme is activated by the detergent Triton X-100, this detergent has no effect on the activities of either serum or acetone powder extract; since high-speed centrifugation does not sediment the enzyme activities of the latter two preparations, it is concluded that they are soluble. Comparison of the membrane-bound and soluble liver enzymes indicates that the membrane modifies kinetic behavior only to a limited extent. In both liver and serum, a single sialyltransferase is responsible for incorporation of sialic acid into α1-acid glycoprotein, fetuin, and N-acetyllactosamine, and sialic acid incorporation occurs whenever a terminal galactose linked (β, 1 → 4) to a penultimate N-acetylglucosamine is presented to the enzyme. Although the serum enzyme resembles the liver enzyme, both the source and function of serum sialyltransferase are unknown.

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Elimination of 2-keto-3-deoxy-D-glycero-D-galacto-nonulosonic acid 9-phosphate synthase activity from human N-acetylneuraminic acid 9-phosphate synthase by a single mutation

Biochemical Journal ◽

10.1042/bj20052034 ◽

2006 ◽

Vol 397 (1) ◽

pp. 195-201 ◽

Cited By ~ 9

Author(s):

Jijun Hao ◽

Willie F. Vann ◽

Stephan Hinderlich ◽

Munirathinam Sundaramoorthy

Keyword(s):

Aldol Condensation ◽

Saturation Mutagenesis ◽

Single Mutation ◽

Wild Type ◽

Wild Type Enzyme ◽

High Sequence Identity ◽

Acetylneuraminic Acid ◽

Nonulosonic Acid ◽

The Impact ◽

Phosphate Synthase

The most commonly occurring sialic acid Neu5Ac (N-acetylneuraminic acid) and its deaminated form, KDN (2-keto-3-deoxy-D-glycero-D-galacto-nonulosonic acid), participate in many biological functions. The human Neu5Ac-9-P (Neu5Ac 9-phosphate) synthase has the unique ability to catalyse the synthesis of not only Neu5Ac-9-P but also KDN-9-P (KDN 9-phosphate). Both reactions are catalysed by the mechanism of aldol condensation of PEP (phosphoenolpyruvate) with sugar substrates, ManNAc-6-P (N-acetylmannosamine 6-phosphate) or Man-6-P (mannose 6-phosphate). Mouse and putative rat Neu5Ac-9-P synthases, however, do not show KDN-9-P synthase activity, despite sharing high sequence identity (>95%) with the human enzyme. Here, we demonstrate that a single mutation, M42T, in human Neu5Ac-9-P synthase can abolish the KDN-9-P synthase activity completely without compromising the Neu5Ac-9-P synthase activity. Saturation mutagenesis of Met42 of the human Neu5Ac-9-P synthase showed that the substitution with all amino acids except leucine retains only the Neu5Ac-9-P synthase activity at levels comparable with the wild-type enzyme. The M42L mutant, like the wild-type enzyme, showed the additional KDN-9-P synthase activity. In the homology model of human Neu5Ac-9-P synthase, Met42 is located 22 Å (1 Å=0.1 nm) away from the substrate-binding site and the impact of this distant residue on the enzyme functions is discussed.

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The Evolutionary Emmergence Of N-Acetylneuraminic Acid (Sialic Acid)- Containing Glycosphingolipids From Deuterosome Echinodermata Starfish

Journal of Glycobiology ◽

10.4172/2168-958x.1000e109 ◽

2014 ◽

Vol 03 (02) ◽

Cited By ~ 1

Author(s):

Cheorl Ho Kim

Keyword(s):

Sialic Acid ◽

Acetylneuraminic Acid

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NontypeableHaemophilus influenzaeHas Evolved Preferential Use ofN-Acetylneuraminic Acid as a Host Adaptation

mBio ◽

10.1128/mbio.00422-19 ◽

2019 ◽

Vol 10 (3) ◽

Cited By ~ 5

Author(s):

Preston S. K. Ng ◽

Christopher J. Day ◽

John M. Atack ◽

Lauren E. Hartley-Tassell ◽

Linda E. Winter ◽

...

Keyword(s):

Sialic Acid ◽

Antibody Response ◽

Immune Evasion ◽

Biofilm Formation ◽

Culture Media ◽

Carbon Sources ◽

Bacterial Pathogen ◽

Catalytic Efficiency ◽

Human Antibody ◽

Acetylneuraminic Acid

ABSTRACTNontypeableHaemophilus inﬂuenzae(NTHi) is a Gram-negative bacterial pathogen that is adapted exclusively to human hosts. NTHi utilizes sialic acid from the host as a carbon source and as a terminal sugar on the outer membrane glycolipid lipooligosaccharide (LOS). Sialic acid expressed on LOS is critical in NTHi biofilm formation and immune evasion. There are two major forms of sialic acids in most mammals,N-acetylneuraminic acid (Neu5Ac) andN-glycolylneuraminic acid (Neu5Gc), the latter of which is derived from Neu5Ac. Humans lack the enzyme to convert Neu5Ac to Neu5Gc and do not express Neu5Gc in normal tissues; instead, Neu5Gc is recognized as a foreign antigen. A recent study showed that dietary Neu5Gc can be acquired by NTHi colonizing humans and then presented on LOS, which acts as an antigen for the initial induction of anti-Neu5Gc antibodies. Here we examined Neu5Gc uptake and presentation on NTHi LOS. We show that, although Neu5Gc and Neu5Ac are utilized equally well as sole carbon sources, Neu5Gc is not incorporated efficiently into LOS. When equal amounts of Neu5Gc and Neu5Ac are provided in culture media, there is ∼4-fold more Neu5Ac incorporated into LOS, suggesting a bias in a step of the LOS biosynthetic pathway. CMP-Neu5Ac synthetase (SiaB) was shown to have ∼4,000-fold-higher catalytic efficiency for Neu5Ac than for Neu5Gc. These data suggest that NTHi has adapted preferential utilization of Neu5Ac, thus avoiding presentation of the nonhuman Neu5Gc in the bacterial cell surface. The selective pressure for this adaptation may represent the human antibody response to the Neu5Gc xenoantigen.IMPORTANCEHost-adapted bacterial pathogens such as NTHi cannot survive out of their host environment and have evolved host-specific mechanisms to obtain nutrients and evade the immune response. Relatively few of these host adaptations have been characterized at the molecular level. NTHi utilizes sialic acid as a nutrient and also incorporates this sugar into LOS, which is important in biofilm formation and immune evasion. In the present study, we showed that NTHi has evolved to preferentially utilize the Neu5Ac form of sialic acid. This adaptation is due to the substrate preference of the enzyme CMP-Neu5Ac synthetase, which synthesizes the activated form of Neu5Ac for macromolecule biosynthesis. This adaptation allows NTHi to evade killing by a human antibody response against the nonhuman sialic acid Neu5Gc.

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