Biochemical characterization of a Neisseria meningitidis polysialyltransferase reveals novel functional motifs in bacterial sialyltransferases

Abteilung Zelluläre Chemie, Medizinische Hochschule Hannover, Carl-Neuberg-Str. 1, 30625 Hannover, Germany.
Molecular Microbiology (Impact Factor: 4.42). 09/2007; 65(5). DOI: 10.1111/j.1365-2958.2007.05862.x
Source: PubMed


The extracellular polysaccharide capsule is an essential virulence factor of Neisseria meningitidis, a leading cause of severe bacterial meningitis and sepsis. Serogroup B strains, the primary disease causing isolates in Europe and America, are encapsulated in alpha-2,8 polysialic acid (polySia). The capsular polymer is synthesized from activated sialic acid by action of a membrane-associated polysialyltransferase (NmB-polyST). Here we present a comprehensive characterization of NmB-polyST. Different from earlier studies, we show that membrane association is not essential for enzyme functionality. Recombinant NmB-polyST was expressed, purified and shown to synthesize long polySia chains in a non-processive manner in vitro. Subsequent structure-function analyses of NmB-polyST based on refined sequence alignments allowed the identification of two functional motifs in bacterial sialyltransferases. Both (D/E-D/E-G and HP motif) are highly conserved among different sialyltransferase families with otherwise little or no sequence identity. Their functional importance for enzyme catalysis and CMP-Neu5Ac binding was demonstrated by mutational analysis of NmB-polyST and is emphasized by structural data available for the Pasteurella multocida sialyltransferase PmST1. Together our data are the first description of conserved functional elements in the highly diverse families of bacterial (poly)sialyltransferases and thus provide an advanced basis for understanding structure-function relations and for phylogenetic sorting of these important enzymes

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    • "The significance for the large conformational changes of the GT-B sialyltransferases is unknown. However, many of these enzymes display substrate promiscuity and alternate activities such as catalyzing the formation of different (a2–3 and a2–6) sialyl linkages [6] [7] [49] [50], or having even sialidase or trans-sialidase activities [6] [7] [8]. Therefore the large conformational changes may be important for these biological functions. "
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    • "In recent years, however, STD NMR studies along with bioinformatics approaches have enabled some insightful structural information [14], though a GT-38 crystal structure remains elusive. All studies done to date indicate that PSTNm requires at least a disialylated oligosaccharide for PST priming [5], and with regard to free sialic acid acceptor, at least a trimer (DP3) is required [15]. Soluble PST–maltose binding protein (PST–MBP) fusions have been expressed, purified, and studied [5]. "
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    • "The known bacterial sialyltransferases do not contain the sialyl motifs found in mammalian sialyltransferases [19,20]. Two short motifs, referred to as the D/E-D/E-G and HP motifs, have been recently identified in the bacterial sialyltransferases and are shown to be functionally important for enzyme catalysis and donor substrate binding [27]. These two motifs are structurally distinct from the motifs found in mammalian sialyltransferases. "
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