CMP-Kdn synthetase catalyses the reaction of sialic acids (Sia) and cytidine-5'-triphosphate (CTP) to the corresponding activated sugar nucleotide CMP-Sia and pyrophosphate PP(i). STD NMR experiments of a recombinant nucleotide cytidine-5'-monophosphate-3-deoxy-d-glycero-d-galacto-nonulosonic acid synthetase (CMP-Kdn synthetase) were performed to map the binding epitope of the substrate CTP and the product CMP-Neu5Ac. The STD NMR analysis clearly shows that the anomeric proton of the ribose moiety of both investigated compounds is in close proximity to the protein surface and is likely to play a key role in the binding process. The relative rates of the enzyme reaction, derived from (1)H NMR signal integrals, show that Kdn is activated at a rate 2.5 and 3.1 faster than Neu5Ac and Neu5Gc, respectively. Furthermore, proton-decoupled (31)P NMR spectroscopy was successfully used to follow the enzyme reaction and clearly confirmed the appearance of CMP-Sia and the inorganic pyrophosphate by-product.
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Identification of a Kdn biosynthesis pathway in the haptophyte suggests widespread sialic acid biosynthesis among microalgae.
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From the Department of Biological Chemistry, John Innes Centre, Norwich Research Park, Norwich NR4 7UH, United Kingdom
Sialic acids are a family of more than 50 structurally distinct acidic sugars on the surface of all vertebrate cells where they terminate glycan chains and are exposed to many interactions with the surrounding environment. In particular, sialic acids play important roles in cell-cell and host-pathogen interactions. The sialic acids or related nonulosonic acids have been observed in Deuterostome lineages, Eubacteria, and Archaea but are notably absent from plants.
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Glycoconj J
July 2006
Institute for Glycomics, Griffith University (Gold Coast Campus), PMB 50 Gold Coast Mail Centre, Queensland 9726, Australia.
CMP-Kdn synthetase catalyses the reaction of sialic acids (Sia) and CTP to the corresponding activated sugar nucleotide CMP-Sia and pyrophosphate PP( i ). Saturation Transfer Difference (STD) NMR spectroscopy has been employed to investigate the sub-structural requirements of the enzyme's binding domain. Sialylnucleoside mimetics, where the sialic acid moiety has been replaced by a carboxyl group and a hydrophobic moiety, have been used in NMR experiments, to probe the tolerance of the CMP-Kdn synthetase to such replacements.
View Article and Find Full Text PDFProbing a CMP-Kdn synthetase by 1H, 31P, and STD NMR spectroscopy.
Biochem Biophys Res Commun
February 2005
Institute for Glycomics, Griffith University (Gold Coast Campus), PMB 60 Gold Coast Mail Centre, Queensland 9726, Australia.
CMP-Kdn synthetase catalyses the reaction of sialic acids (Sia) and cytidine-5'-triphosphate (CTP) to the corresponding activated sugar nucleotide CMP-Sia and pyrophosphate PP(i). STD NMR experiments of a recombinant nucleotide cytidine-5'-monophosphate-3-deoxy-d-glycero-d-galacto-nonulosonic acid synthetase (CMP-Kdn synthetase) were performed to map the binding epitope of the substrate CTP and the product CMP-Neu5Ac. The STD NMR analysis clearly shows that the anomeric proton of the ribose moiety of both investigated compounds is in close proximity to the protein surface and is likely to play a key role in the binding process.
View Article and Find Full Text PDF[KDNology: a new frontier in glycobiology].
Tanpakushitsu Kakusan Koso
December 1998
Institute of Biological Chemistry, Academia Sinica, Taipei, Taiwan, R.O.C.
Substrate specificity of rainbow trout testis CMP-3-deoxy-D-glycero-D-galacto-nonulosonic acid (CMP-Kdn) synthetase: kinetic studies of the reaction of natural and synthetic analogues of nonulosonic acid catalyzed by CMP-Kdn synthetase.
Eur J Biochem
March 1996
Department of Biophysics and Biochemistry, Graduate School of Science, University of Tokyo, Japan.
In this report we present kinetic data of the activation reaction of several synthetic 3-deoxy-D-glycero-D-galacto-nonulosonic acid (Kdn) and N-acetylneuraminic acid (Neu5Ac) analogues catalyzed by the rainbow trout testis CMP-Kdn synthetase. This enzyme showed broad substrate specificity in terms of substitutions at C4 or C5 position of Kdn and Neu5Ac. In contrast, calf brain CMP-N-acylneuraminic acid synthetase had narrow substrate specificity, being active only on various N-acyl analogues of Neu5Ac and only slightly active on Kdn derivatives.
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