Studies on the Substrate Specificity of Acylneuraminate Pyruvate-Lyase

Schauer, Roland; Wember, Margret; Wirtz-Peitz, Ferdinand; Amaral, Clarice Ferreira Do

doi:10.1515/bchm2.1971.352.2.1073

Cited by 50 publications

(22 citation statements)

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“…However, the original study using the E. coli deletion strains indicated that the nature of the C-5 amino substituent in Sia does not affect transport or degradation (60). The aldolases from C. perfringens and E. coli are capable of cleaving a range of neuraminic acid derivatives with different substituents at C-5, including formyl, succinyl, and glycolyl neuraminic acids (1,48). Regarding the NanT permease, it has been established that many bacteria, both gram negative and gram positive, exhibit an active proton symporter-type mechanism (59).…”

Section: Resultsmentioning

confidence: 99%

Crystal Structure of the Bacterial YhcH Protein Indicates a Role in Sialic Acid Catabolism

et al. 2005

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The yhcH gene is part of the nan operon in bacteria that encodes proteins involved in sialic acid catabolism. Determination of the crystal structure of YhcH from Haemophilus influenzae was undertaken as part of a structural genomics effort in order to assist with the functional assignment of the protein. The structure was determined at 2.2-Å resolution by multiple-wavelength anomalous diffraction. The protein fold is a variation of the double-stranded ␤-helix. Two antiparallel ␤-sheets form a funnel opened at one side, where a putative active site contains a copper ion coordinated to the side chains of two histidine and two carboxylic acid residues. A comparison to other proteins with a similar fold and analysis of the genomic context suggested that YhcH may be a sugar isomerase involved in processing of exogenous sialic acid.

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Section: Resultsmentioning

confidence: 99%

Crystal Structure of the Bacterial YhcH Protein Indicates a Role in Sialic Acid Catabolism

et al. 2005

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“…Activation of 9-substituted sialic acid analogues was studied using a partially purified synthase preparation from bovine brain [29,35,361. A preparation of this enzyme, suitable for a large-scale synthesis of CMP-NeuAc, has been already published by van Dijk and van den Eijnden [37].…”

Section: Discussionmentioning

confidence: 99%

Activation and transfer of novel synthetic 9‐substituted sialic acids

Gross

Bünsch

Paulson

et al. 1987

European Journal of Biochemistry

108

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In this report several NeuAc analogues differently modified at position C-9 were tested as substrates for CMPsialic acid synthase from bovine brain: the hydroxy group at C-9 was replaced by an amino, acetamido, benzamido, hexanoylamido and azido group.The synthase was partially purified by chromatography on CDP-hexanolamine -Sepharose. CMP-glycosides synthesized were measured by analytical HPLC at 275 nm.Each NeuAc analogue was activated to the respective CMP-glycoside: K,-values varied from 0.8 mM to 4.6 mM, the K,,, for NeuAc was 1.4 mM. Thus affinity of the enzyme was influenced only moderately by chemical modification at C-9.CMP-glycosides were synthesized on a preparative scale with good yield and characterized by analytical HPLC. in addition, 500-MHz 'H-NMR data of CMP-9-amino-NeuAc and CMP-9-acetamido-NeuAc were obtained.Each CMP-activated NeuAc analogue was a suitable donor substrate for GalP 1-4GlcNAc cr2,6-sialyltransferase from rat liver. Transfer was determined by the thiobarbituric acid method and by analytical HPLC at 200 nm. The results demonstrate that synthetic, not naturally occurring, non-labelled NeuAc analogues can be incorporated into glycoprotein with high yield. Abbreviations. NeuAc, N-acetyl-D-neuraminic acid; 9-aminoNeuAc, 5-acetamido-9-amino-3,5,9-trideoxy-2-nonulosonic acid; 9-acetamido-NeuAc, 5,9-diacetamido-3,5,9-trideoxy-2-nonulosonic acid; 9-hexanoylamido-NeuAc, 5-acetamido-3,5,9-trideoxy-9-hexanoylamido-2-nonulosonic acid; 9-benzamido-NeuAc,5-acetamido-9-benzamido-3,5,9-trideoxy-2-nonulosonic acid; 9-azido-NeuAc, 5-acetamido-9-azido-3,5,9-trideoxy-2-nonulosonic acid ; 9-iodo-NeuAc, 5-acetamido-3,5,9-trideoxy-9-iodo-2-nonulosonic acid; 9-fluoroNeuAc, 5-acetamido-3,5,9-trideoxy-9-fluoro-2-nonulosonic acid; C,-NeuAc, 5-acetamido-3,5-dideoxy-2-heptulosonic acid; CMPNeuAc, cytidine-5'-monophospho-N-acetylneuraminic acid; 9-0-acetyl-NeuAc, 5-acetamido-9-O-acetyl-3,5-dideoxy-2-nonulosonic acid.Enzymes. CMPsialic acid synthase (EC 2.7.7.43); Galpl4GlcNAc a-2,6-sialyltransferase (EC 2.4.99.1); sialidase, acylneuraminylhydrolase (EC 3.2.1.18). cells may be an important biological role of sialic acids [4, 91. But many questions concerning the biological significance of this acid still remain to be answered. Further insight into these problems could be gained by studying synthetic sialic acid analogues. A substitution of natural sialic acid of glycoproteins and glycolipids with synthetic analogues would be of particular interest as the biological role of sialic acids and the correlation between structure and function of sialic acid can be investigated. In principle this could be done enzymatically by removing sialic acid with sialidase, followed by replacement with sialyltransferase and the appropriate donor substrate. In order to carry out such investigations the substrate specificity of CMP sialic acid synthase and sialyltransferase for each NeuAc analogue has to be established first.To date, only a few synthetic analogues have been used as substrate for the synthase. Although the activati...

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“…The enzyme was assayed by the L-lactate dehydrogenase method according to [7]. The concentrations of NeuAc and Me4NeuAc were in the range of 0.2 to 2 mM.…”

Section: Assuj~ For Acylneuraminate Pyruvale-lyusementioning

confidence: 99%

“…It has been described that the 4-0-acetyl-Nacetyl-(or -N-glycoloy1)neuraminic acid, when bound to glycoconjugates, is not cleaved by bacterial [5] or mammalian [6] neuraminidases and that free 4-0-acetyl-N-acetylneuraminic acid is largely resistant to degradation by the acylneuraminate pyruvate-lyase (aldolase) [7].…”

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Metabolism of 4‐O‐Methyl‐N‐acetylneuraminic Acid a Synthetic Sialic Acid

Beau

Schauer

1980

European Journal of Biochemistry

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1. 4-0-Methyl-N-acetylneuraminic acid shows a strong positive periodate-thiobarbiturate reaction. The mechanism of dye formation in this test for sialic acids is discussed in view of the studies already published.2. An efficient preparation of a tritium-labelled 4-0-methyl-N-acetylneuraminic acid, with high specific radioactivity, by an oxymercuration-demercuration procedure is presented.3. Sialyltransferase activities in microsomal fractions of equine liver using desialylated fetuin are studied. The enzyme activity, assayed in a radioactive procedure, shows an apparent K , value for CMP-N-acetylneuraminic acid of 0.7 mM, whereas this value is 3.4 mM for CMP-4-0-methyl-N-acetylneuraminic acid. Differences are also observed in the maximal velocity for the two substrates.4. The equine liver system can be used to prepare substantial amounts of fetuin containing radioactive N-acetylneuraminic acid or 4-0-methyl-N-acetylneuraminic acid. The isolated reaction products show similar sialic acid release by treatments with acid or fowl-plague virus neuraminidase. In contrast, 4-0-methyl-N-acetylneuraminic acid-fetuin displays a marked resistance to desialylation by Vibrio cholerae neuraminidase.5. Free 4-0-methyl-N-acetylneuraminic acid is completely resistant to the action of acylneuraminate pyruvate-lyase. It does not inhibit the enzymic cleavage reaction of N-acetylneuraminic acid.6. The influence of a substitution at C-4 of neuraminic acid on the enzymatic reaction mechanisms is discussed.The acylneuraminic acids (sialic acids), widely distributed in animal tissues, are important components of oligosaccharides, glycolipids and glycoproteins. The occurrence and nature of various Nacylated and O-acylated neuraminic acids is now well documented [l -31, and the pattern distribution is characteristic for the animal species. Sialic acids are implicated in many cellular processes [4], and the high number of the 0-acylated derivatives points to the importance of understanding the biological role of these substances. In a preliminary approach, a precise evaluation of the behaviour of the main enzymes involved in the metabolism of sialic acids is

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Studies on the Substrate Specificity of Acylneuraminate Pyruvate-Lyase

Cited by 50 publications

References 1 publication

Crystal Structure of the Bacterial YhcH Protein Indicates a Role in Sialic Acid Catabolism

Crystal Structure of the Bacterial YhcH Protein Indicates a Role in Sialic Acid Catabolism

Activation and transfer of novel synthetic 9‐substituted sialic acids

Metabolism of 4‐O‐Methyl‐N‐acetylneuraminic Acid a Synthetic Sialic Acid

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