CMP-Sialic Acid Synthetase: The Point of Constriction in the Sialylation Pathway

Sellmeier, Melanie; Weinhold, Birgit; Münster-Kühnel, Anja K.

doi:10.1007/128_2013_477

Cited by 24 publications

(30 citation statements)

References 132 publications

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“…Previous studies revealed that divalent metals, such as Mg 2+ or Mn 2+ , are essential for the activity CMP-Sia synthetases as these metal ions bind in the vicinity of the enzyme active site and facilitate the orientation of substrates and catalysis [4]. Mg 2+ is thought to be the preferred metal cofactor for CMP-Sia synthetases, and it was used in the majority of previous studies on in vitro activities of the synthetases.…”

Section: Resultsmentioning

confidence: 99%

“…The presence of divalent metal cations is a strict requirement for the activity of CMP-Sia synthetase [4], including Drosophila CSAS (Fig. 4A).…”

Section: Discussionmentioning

confidence: 99%

“…Only N-terminal parts of sequences corresponding to the CMP-Sia synthetase domain were used for enzymes with bipartite domain architecture [4]. Phylogenetic tree was assembled by ClustalW2 program using neighbor-joining algorithm [18].…”

Section: Methodsmentioning

confidence: 99%

“…While organisms can possess multiple sialyltransferases (e.g. humans have twenty different sialyltransferase enzymes with distinct linkage and substrate specificities), there is usually only one CSAS (aka CMAS) enzyme mediating the sialic acid activation step in all known organisms mediating de novo sialylation, with the exception of some fish species that have two CSAS genes due to a presumptive gene duplication [4]. …”

Section: Introductionmentioning

confidence: 99%

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Characterization of Drosophila CMP-sialic acid synthetase activity reveals unusual enzymatic properties

Мерцалов

Novikov

Scott

et al. 2016

Biochemical Journal

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CMP-sialic acid synthetase (CSAS) is a key enzyme of the sialylation pathway. CSAS produces the activated sugar donor, CMP-sialic acid, which serves as a substrate for sialyltransferases to modify glycan termini with sialic acid. Unlike other animal CMP-Sia synthetases that normally localize in the nucleus, Drosophila melanogaster CSAS (DmCSAS) localizes in the cell secretory compartment, predominantly in the Golgi, which suggests that this enzyme has properties distinct from those of its vertebrate counterparts. To test this hypothesis, we purified recombinant DmCSAS and characterised its activity in vitro. Our experiments revealed several unique features of this enzyme. DmCSAS displays specificity for N-acetylneuraminic acid as a substrate, shows preference for lower pH and can function with a broad range of metal cofactors. When tested at a pH corresponding to the Golgi compartment, the enzyme showed significant activity with several metal cations, including Zn2+, Fe2+, Co2+ and Mn2+, while the activity with Mg2+ was found to be low. Protein sequence analysis and site-specific mutagenesis identified an aspartic acid residue that is necessary for enzymatic activity and predicted to be involved in coordinating a metal cofactor. DmCSAS enzymatic activity was found to be essential in vivo for rescuing the phenotype of DmCSAS mutants. Finally, our experiments revealed a steep dependence of the enzymatic activity on temperature. Taken together, our results indicate that DmCSAS underwent evolutionary adaptation to pH and ionic environment different from that of counterpart synthetases in vertebrates. Our data also suggest that environmental temperatures can regulate Drosophila sialylation, thus modulating neural transmission.

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Section: Resultsmentioning

confidence: 99%

“…The presence of divalent metal cations is a strict requirement for the activity of CMP-Sia synthetase [4], including Drosophila CSAS (Fig. 4A).…”

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Characterization of Drosophila CMP-sialic acid synthetase activity reveals unusual enzymatic properties

Мерцалов

Novikov

Scott

et al. 2016

Biochemical Journal

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show abstract

“…In animals and microbes, sialic acid must be activated for use in glycan biosynthesis by conjugation with CTP, a process catalyzed by CMP-Sialic acid synthetase (Kean et al, 2004). These are encoded by the CMAS gene, which is highly conserved among vertebrates and well-conserved even in other prokaryotes and eukaryotes (Sellmeier et al, 2013). No genes with significant homology spanning the functional domains of this gene were found in C. elegans or Schistosoma genomes.…”

Section: Glycan Motifsmentioning

confidence: 99%

Deciphering the glycogenome of schistosomes

et al. 2014

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Schistosoma mansoni and other Schistosoma sp. are multicellular parasitic helminths (worms) that infect humans and mammals worldwide. Infection by these parasites, which results in developmental maturation and sexual differentiation of the worms over a period of 5–6 weeks, induces antibodies to glycan antigens expressed in surface and secreted glycoproteins and glycolipids. There is growing interest in defining these unusual parasite-synthesized glycan antigens and using them to understand immune responses, their roles in immunomodulation, and in using glycan antigens as potential vaccine targets. A key problem in this area, however, has been the lack of information about the enzymes involved in elaborating the complex repertoire of glycans represented by the schistosome glycome. Recent availability of the nuclear genome sequences for Schistosoma sp. has created the opportunity to define the glycogenome, which represents the specific genes and cognate enzymes that generate the glycome. Here we describe the current state of information in regard to the schistosome glycogenome and glycome and highlight the important classes of glycans and glycogenes that may be important in their generation.

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A Microfluidic Countercurrent Reactor for Accelerating Enzymatic Reactions

et al. 2023

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Enzymes are important catalysts in biochemical reactions with superior regio, stereo, and substrate selectivity. However, enzymatic reaction systems have drawbacks including product inhibition, difficulty recycling, and poor stability. Importantly, the rate of an enzyme catalyzed reaction diminishes rapidly due to product inhibition and substrate depletion, making it difficult for many enzymes to catalyze a reaction to completion. The outcome is a mixture of unreacted substrates being present in the final reaction, necessitating additional separation steps that increase costs. This study presents a microfluidic reactor for accelerating enzyme catalyzed reactions using a countercurrent design that continuously removes products and adds fresh substrate into the reaction, allowing enzymes to operate under better reaction conditions. It demonstrates that countercurrent flow accelerates enzymatic reactions in our system up to 36 % for horseradish peroxidase and 21 % for β‐glucosidase compared to cocurrent flow, and the resulting reaction solution contains highly pure product with minimal substrate contamination.

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CMP-Sialic Acid Synthetase: The Point of Constriction in the Sialylation Pathway

Cited by 24 publications

References 132 publications

Characterization of Drosophila CMP-sialic acid synthetase activity reveals unusual enzymatic properties

Characterization of Drosophila CMP-sialic acid synthetase activity reveals unusual enzymatic properties

Deciphering the glycogenome of schistosomes

A Microfluidic Countercurrent Reactor for Accelerating Enzymatic Reactions

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