Functional and structural studies on the Neisseria gonorrhoeae GmhA, the first enzyme in the glycero-manno-heptose biosynthesis pathways, demonstrate a critical role in lipooligosaccharide synthesis and gonococcal viability

Igor H. Wierzbicki, Ryszard A. Zielke, Konstantin V. Korotkov, Aleksandra E. Sikora

Research output: Contribution to journalArticlepeer-review

18 Scopus citations

Abstract

Sedoheptulose-7-phosphate isomerase, GmhA, is the first enzyme in the biosynthesis of nucleotide-activated-glycero-manno-heptoses and an attractive, yet underexploited, target for development of broad-spectrum antibiotics. We demonstrated that GmhA homologs in Neisseria gonorrhoeae and N. meningitidis (hereafter called GmhAGC and GmhANM, respectively) were interchangeable proteins essential for lipooligosaccharide (LOS) synthesis, and their depletion had adverse effects on neisserial viability. In contrast, the Escherichia coli ortholog failed to complement GmhAGC depletion. Furthermore, we showed that GmhAGC is a cytoplasmic enzyme with induced expression at mid-logarithmic phase, upon iron deprivation and anaerobiosis, and conserved in contemporary gonococcal clinical isolates including the 2016 WHO reference strains. The untagged GmhAGC crystallized as a tetramer in the closed conformation with four zinc ions in the active site, supporting that this is most likely the catalytically active conformation of the enzyme. Finally, site-directed mutagenesis studies showed that the active site residues E65 and H183 were important for LOS synthesis but not for GmhAGC function in bacterial viability. Our studies bring insights into the importance and mechanism of action of GmhA and may ultimately facilitate targeting the enzyme with small molecule inhibitors.

Original languageEnglish
Article numbere00432
JournalMicrobiologyOpen
Volume6
Issue number2
DOIs
StatePublished - Apr 1 2017

Bibliographical note

Publisher Copyright:
© 2017 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

Keywords

  • Neisseria gonorrhoeae
  • crystal structure
  • drug target
  • sedoheptulose-7-phosphate isomerase GmhA

ASJC Scopus subject areas

  • Microbiology

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