Lipooligosaccharides in the outer membrane of pathogenic Haemophilus influenzae and Haemophilus ducreyi species have been found to contain sialic acid. It is believed thatterminal lactosamine (Galb1'4GlcNAc) is the acceptor for sialic acid. In the related mucosal pathogens Neisseria gonorrhoeae and N. meningitidis, sialylation of LOS has been shown to haveimportant implications in the origanisms' ability to evade lysis by human serum and killing by human neutrophils. We, therefore, propose to isolate and characterize the sialyltransferase fromH. ducreyi and to develop inhibitors of this enzyme as a potential therapeutic. We plan to use mass spectrometry to aid our initial isolations and sequence analysis of this sialyltransferase (ST). Laser desorption and electrospray mass spectrometry will be used to provide accurate molecular weight information and help establish the purity of the protein. Tandem mass spectrometry will then be used to sequence regions of the sialyltransferase with the aim of constructing oligonucleotide probes to isolate the cDNA encoding this enzyme. Lastly, conventional mass spectrometric techniques such as electron impact (EI) and chemical ionization (CI)mass spectrometry will help our efforts to synthesize novel inhibitors based on the required substrates of this enzyme, CMP-NANA and/or the acceptor disaccharide, lactosamine.
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