SummaryNeisseria gonorrhoeae expresses an O-linked protein glycosylation pathway that targets PilE, the major pilin subunit protein of the Type IV pilus colonization factor. Efforts to define glycan structure and thus the functions of pilin glycosylation (Pgl) components at the molecular level have been hindered by the lack of sensitive methodologies. Here, we utilized a 'topdown' mass spectrometric approach to characterize glycan status using intact pilin protein from isogenic mutants. These structural data enabled us to directly infer the function of six components required for pilin glycosylation and to define the glycan repertoire of strain N400. Additionally, we found that the N. gonorrhoeae pilin glycan is O-acetylated, and identified an enzyme essential for this unique modification. We also identified the N. gonorrhoeae pilin oligosaccharyltransferase using bioinformatics and confirmed its role in pilin glycosylation by directed mutagenesis. Finally, we examined the effects of expressing the PglA glycosyltransferase from the Campylobacter jejuni N-linked glycosylation system that adds N-acetylgalactosamine onto undecaprenylpyrophosphate-linked bacillosamine. The results indicate that the C. jejuni and N. gonorrhoeae pathways can interact in the synthesis of O-linked di-and trisaccharides, and therefore provide the first experimental evidence that biosynthesis of the N. gonorrhoeae pilin glycan involves a lipid-linked oligosaccharide precursor. Together, these findings underpin more detailed studies of pilin glycosylation biology in both N. gonorrhoeae and N. meningitidis, and demonstrate how components of bacterial O-and N-linked pathways can be combined in novel glycoengineering strategies.

Dimethyl disulfide (DMDS) and N-methylacetamide are two first choice model systems that represent the disulfide bridge bonding and the peptide bonding in proteins. These molecules are therefore suitable for investigation of the mechanisms involved when proteins fragment under electron capture dissociation (ECD). The dissociative recombination cross sections for both protonated DMDS and protonated N-methylacetamide were determined at electron energies ranging from 0.001 to 0.3 eV. Also, the branching ratios at 0 eV center-of-mass collision energy were determined. The present results give support for the indirect mechanism of ECD, where free hydrogen atoms produced in the initial fragmentation step induce further decomposition. We suggest that both indirect and direct dissociations play a role in ECD.

A simple hydride‐transfer mechanism to produce alkyl cations occurs when isolated protonated hydrogen peroxide (HOOH2+) attacks the C−H bonds of alkanes (except methane) [Eq. (1)]. The chemical properties of HOOH2+, which may serve as a model of the central catalytic motif of key biological systems, have been studied for the first time.

Abstract. Atomic emission spectroscopy coupled with capillary gas chromatography (GC) has been evaluated for molecular formula determination of several chlorinated and brominated compounds present in three environmental samples. Variations in elemental responses due to compound type, poor chromatographic resolution, and small signals relative to the background were all found to complicate molecular formula calculations. About 20% of the calculated molecular formulas presented in this paper agreed with theoretical values. For most applications, identifications based solely on GC coupled with atomic emission detection (AED) are not sufficiently accurate. However, the combination of elemental information derived from a GC-AED analysis with complementary data obtained from other detection techniques results in a strong analytical tool for analyte identification.

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