Bacterial CMP-sialic acid synthetases: Production, properties, and applications
ABSTRACT Sialic acids are abundant nine-carbon sugars expressed terminally on glycoconjugates of eukaryotic cells and are crucial for a variety of cell biological functions such as cell-cell adhesion, intracellular signaling, and in regulation of glycoproteins stability. In bacteria, N-acetylneuraminic acid (Neu5Ac) polymers are important virulence factors. Cytidine 5'-monophosphate (CMP)-N-acetylneuraminic acid synthetase (CSS; EC 18.104.22.168), the key enzyme that synthesizes CMP-N-acetylneuraminic acid, the donor molecule for numerous sialyltransferase reactions, is present in both prokaryotes and eukaryotic systems. Herein, we emphasize the source, function, and biotechnological applications of CSS enzymes from bacterial sources. To date, only a few CSS from pathogenic bacterial species such as Neisseria meningitidis, Escherichia coli, group B streptococci, Haemophilus ducreyi, and Pasteurella hemolytica and an enzyme from nonpathogenic bacterium, Clostridium thermocellum, have been described. Overall, the enzymes from both Gram-positive and Gram-negative bacteria share common catalytic properties such as their dependency on divalent cation, temperature and pH profiles, and catalytic mechanisms. The enzymes, however, can be categorized as smaller and larger enzymes depending on their molecular weight. The larger enzymes in some cases are bifunctional; they have exhibited acetylhydrolase activity in addition to their sugar nucleotidyltransferase activity. The CSSs are important enzymes for the chemoenzymatic synthesis of various sialooligosaccharides of significance in biotechnology.
- SourceAvailable from: Yanhong Li
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- "The crystal structures of NmCSS (Horsfall et al. 2010) and N-terminal catalytically active domain of murine CSS (Krapp et al. 2003) have been reported. The enzyme has also been cloned from E. coli K1 (Yu et al. 2004), S. agalactiae or GBS (Yu et al. 2006c), H. ducreyi (Tullius et al. 1996), Pasteurella haemolytica A2 (Bravo et al. 2001), Clostridium thermocellum (Mizanur and Pohl 2007) as summarized in a recent review (Mizanur and Pohl 2008) and more recently from P. multocida (Li et al. 2012). The substrate promiscuity of NmCSS enables its application in catalyzing the formation of diverse CMP-sialic acids (Morley and Withers 2010; Yu et al. 2004; Rauvolfova et al. 2008; Hartlieb et al. 2008; Gilbert et al. 1998). "
ABSTRACT: Sialic acids are a family of negatively charged monosaccharides which are commonly presented as the terminal residues in glycans of the glycoconjugates on eukaryotic cell surface or as components of capsular polysaccharides or lipooligosaccharides of some pathogenic bacteria. Due to their important biological and pathological functions, the biosynthesis, activation, transfer, breaking down, and recycle of sialic acids are attracting increasing attention. The understanding of the sialic acid metabolism in eukaryotes and bacteria leads to the development of metabolic engineering approaches for elucidating the important functions of sialic acid in mammalian systems and for large-scale production of sialosides using engineered bacterial cells. As the key enzymes in biosynthesis of sialylated structures, sialyltransferases have been continuously identified from various sources and characterized. Protein crystal structures of seven sialyltransferases have been reported. Wild-type sialyltransferases and their mutants have been applied with or without other sialoside biosynthetic enzymes for producing complex sialic acid-containing oligosaccharides and glycoconjugates. This mini-review focuses on current understanding and applications of sialic acid metabolism and sialyltransferases.Applied Microbiology and Biotechnology 04/2012; 94(4):887-905. DOI:10.1007/s00253-012-4040-1 · 3.81 Impact Factor
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- "Some bacteria produce PSA as their capsules, such as Neisseria meningitidis, Escherichia coli (E. coli), group B Streptococci, Haemophilus ducreyi and Pasteurella hemolytica (Mizanur and Pohl, 2008). In E. coli K1, the PSA capsule consists of linear homo ␣2,8-linked N-acetylneuraminic acids (Neu5Ac) with the degree of polymerization up to 200 (Pelkonen et al., 1988). "
ABSTRACT: The polysialic acid (PSA) production in Escherichia coli (E. coli) K1 was studied using three different cultivation strategies. A batch cultivation, a fed-batch cultivation at a constant specific growth rate of 0.25 h(-1) and a fed-batch cultivation at a constant glucose concentration of 50 mg l(-1) was performed. PSA formation kinetics under different cultivation strategies were analyzed based on the Monod growth model and the Luedeking-Piret equation. The results revealed that PSA formation in E. coli K1 was completely growth associated, the highest specific PSA formation rate (0.0489 g g(-1)h(-1)) was obtained in the batch cultivation. However, comparing biomass and PSA yields on the glucose consumed, both fed-batch cultivations provided higher yields than that of the batch cultivation and acetate formation was prevented. Moreover, PSA yield on glucose was also correlated to the specific growth rate of the cells. The optimal specific growth rate for PSA production was 0.32 h(-1) obtained in the fed-batch cultivation at a constant glucose concentration of 50 mg l(-1), with highest conversion efficiency of 43 mg g(-1).Journal of Biotechnology 04/2011; 154(4):222-9. DOI:10.1016/j.jbiotec.2011.04.009 · 2.88 Impact Factor
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- "Due to limited availability, the market price of PSA is as high as US$200 per gram . Bacteria such as Neisseria meningitides, Escherichia coli, Haemophilus ducreyi and Pasteurella haemolytica have been found to produce capsular PSA in their culture broth  . "
ABSTRACT: Polysialic acid (PSA) is a novel pharmaceutical material used in control release for protein drugs or in biomedical applications as scaffold. An efficient pilot production process for bacterial PSA was developed. Our PSA fermentation process by Escherichia coli CCTCC M208088 was optimized in a 500L fermenter using a novel strategy by controlling pH with ammonia water feeding coupled with sorbitol supplementation. The resulting PSA level increased to 5500mg/L as compared with the 1500mg/L of the control. Furthermore, the process for the PSA purification from the fermentation broth was also established. PSA was isolated from the broth by ethanol precipitation, filtration with perlite as filter aid, followed by cetyl pyridinium chloride (CPC) precipitation and lyophilization. The final PSA product obtained had 98.1±1.6% purity at 56.1±1.7% recovery rate. Infrared spectroscopy and NMR spectroscopy analysis indicated that the structure of resulting PSA was identical to the published α-2,8 linked polysialic acid.Biochemical Engineering Journal 12/2010; 53(1):97-103. DOI:10.1016/j.bej.2010.09.015 · 2.37 Impact Factor