Scientific Publications by FDA Staff

J Bacteriol 2011 Apr;193(7):1576-82

Characterization and acceptor preference of a soluble meningococcal group C polysialyltransferase.

Peterson DC, Arakere G, Vionnet J, McCarthy P, Vann WF

Vaccines against Neisseria meningitidis (Nm) group C are based on its ¿2,9 linked polysialic acid capsular polysaccharide. This polysialic acid expressed on the surface of N. meningitidis and in the absence of specific antibody, serves to evade host defense mechanisms (1). The polysialyltransferase that forms the group C polysialic acid is located in the cytoplasmic membrane. Until recently detailed characterization of bacterial polysialyltransferases has been hampered by a lack of availability of soluble enzyme preparations (2, 3). We have constructed chimera of the group C polysialyltransferase that catalyze the formation ¿2,9 polysialic acid as a soluble enzyme. We have used site directed mutagenesis to determine the region of the enzyme necessary for synthesis of the ¿2,9 linkage. A chimera of NmB and NmC polysialyltransferase (PST) containing only amino acids 1-107 of the NmB polysialyltransferase catalyzed the synthesis of ¿2,8 polysialic acid. The NmC polysialyltransferase requires an exogenous acceptor for catalytic activity. While it requires a minimum of a disialylated oligosaccharide to catalyze transfer, it can form high MW ¿2,9 polysialic acid in a non-processive fashion when initiated with an ¿2,8 polysialic acid acceptor. De novo synthesis in vivo requires an endogenous acceptor. We attempted to reconstitute de novo activity of the soluble group C polysialyltransferase with membrane components. We show that an acapsular mutant with a defect in the polysialyltransferase produces outer membrane vesicles containing an acceptor for the ¿2,9 polysialyltransferase. This acceptor is an amphipathic molecule and can be elongated to produce polysialic acid reactive with group C specific antibody.