Chemoenzymatic synthesis of C8-modified sialic acids and related α2–3- and α2–6-linked sialosides

Abstract

Naturally occurring 8-O-methylated sialic acids, including 8-O-methyl-N-acetylneuraminic acid and 8-O-methyl-N-glycolylneuraminic acid, along with 8-O-methyl-2-keto-3-deoxy-D-glycero-D-galacto-nonulosonic acid (Kdn8Me) and 8-deoxy-Kdn were synthesized from corresponding 5-O-modified six-carbon monosaccharides and pyruvate using a sialic acid aldolase cloned from Pasteurella multocida strain P-1059 (PmNanA). In addition, α2–3- and α2–6-linked sialyltrisaccharides containing Neu5Ac8Me and Kdn8Deoxy were also synthesized using a one-pot multienzyme approach. The strategy reported here provides an efficient approach to produce glycans containing various C8-modified sialic acids for biological evaluations.

Sialic acid metabolism and sialyltransferases: natural functions and applications

“…Both C2- and C5-modified N -acetylmannosamine (ManNAc) or mannose derivatives can be used as substrates by E. coli K-12 sialic acid aldolase (EcNanA) and P. multocida P-1059 sialic acid aldolase (PmNanA) (Li et al 2008; Cao et al 2009b). Nevertheless, PmNanA is a more efficient enzyme than EcNanA for synthesizing C8-modi ed sialic acids, especially for 5- O -methyl ManNAc, 5- O -methyl N -glycolylmannosamine (ManNGc5OMe), 5- O -methyl mannose, and 5-deoxy-mannose (Yu et al 2011; Li et al 2008). For activating sialic acid to form CMP-sialic acid, the donor of sialyltransferase, CMP-sialic acid synthetase (CSS, or sialic acid cytidylyltransferase, EC 2.7.7.43) from N. meningitidis (NmCSS) was the first bacterial CSS that was characterized in detail (Warren and Blacklow 1962; Yu et al 2004).…”

“…Using the one-pot three-enzyme sialylation system containing a multifunctional P. multocida sialyltransferase PmST1 encoded by Pm0188 gene, an E. coli or a P. multocida sialic acid aldolase (EcNanA or PmNanA), and NmCSS, α2–3-linked structurally diverse sialosides containing C5-, C8-, C9-, and other modified sialic acids and/or various acceptors including mono- and oligosaccharides, were synthesized in preparative-scale at 37°C, pH 8.5 (for sialosides that do not have a base labile O -acetyl or O -lactyl group) or pH 7.5 (for sialosides with an O -acetyl or O -lactyl group) (Yu et al 2005; Yu et al 2011; Cao et al 2008; Lau et al 2011; Yu et al 2006a). Typical yields for preparative-scale (>20 mg) synthesis were higher than 60%, many reactions were achieved with more than 90% yields.…”

“…The tumor-associated sialyl T-antigens and derivatives were also synthesized using PmST1 in an efficient sequential two-step multienzyme approach (Lau et al 2011). Recently, both PmST1 and Cst-I, an α2–3-sialyltransferase from C. jejuni , have been used synthesizing sialosides containing C8-modified sialic acid (Yu et al 2011; Morley and Withers 2010). …”

“…The extremely flexible substrate specificity of the P. damselae α2–6 sialyltransferase (Pd2,6ST) enables its application in highly efficient chemoenzymatic synthesis of naturally occurring and non-naturally α2–6-linked sialosides (Yu et al 2006b; Yu et al 2011; Yu et al 2006a; Muthana et al 2007). Size-defined polysaccharide analogues were also chemoenzymatically synthesis by Pd2,6ST-catalyzed block transfer of di- or tetra-oligosaccharides from their CMP-activated forms (Muthana et al 2007).…”

Chemoenzymatic synthesis of sialosides containing C7-modified sialic acids and their application in sialidase substrate specificity studies

“…Sialosides with natural and non-natural modifications at C-5, C-9, and C-8 of sialic acid have been successfully synthesized. 6–12 α2–3- and α2–6-Linked sialosides Siaα2–3/6GalβpNP containing C-5 and/or C-9 modified sialic acids have been used to elucidate the substrate specificities of bacterial, human, and viral sialidases. 9–11,13 Selective sialidase inhibitors against human NEU2 or bacterial sialidases have been designed and synthesized based on the structural features of the sialic acid modification obtained from sialidase substrate specificity studies.…”