By the same authors

From the same journal

From the same journal

The basis for non-canonical ROK family function in the N-acetylmannosamine kinase from the pathogen Staphylococcus aureus

Research output: Contribution to journalArticlepeer-review

Full text download(s)

Published copy (DOI)


  • David Coombes
  • James S Davies
  • Michael C Newton-Vesty
  • Christopher R Horne
  • Thanuja G Setty
  • Ramaswamy Subramanian
  • James W B Moir
  • Rosmarie Friemann
  • Santosh Panjikar
  • Michael D W Griffin
  • Rachel A North
  • Renwick C J Dobson


Publication details

JournalThe Journal of biological chemistry
DateAccepted/In press - 15 Jan 2020
DateE-pub ahead of print - 15 Jan 2020
DatePublished (current) - 6 Mar 2020
Issue number10
Number of pages15
Pages (from-to)3301-3315
Early online date15/01/20
Original languageEnglish


In environments where glucose is limited, some pathogenic bacteria metabolize host-derived sialic acid as a nutrient source. N-Acetylmannosamine kinase (NanK) is the second enzyme of the bacterial sialic acid import and degradation pathway and adds phosphate to N-acetylmannosamine using ATP to prime the molecule for future pathway reactions. Sequence alignments reveal that Gram-positive NanK enzymes belong to the Repressor, ORF, Kinase (ROK) family, but many lack the canonical Zn-binding motif expected for this function, and the sugar-binding EXGH motif is altered to EXGY. As a result, it is unclear how they perform this important reaction. Here, we study the Staphylococcus aureus NanK (SaNanK), which is the first characterization of a Gram-positive NanK. We report the kinetic activity of SaNanK along with the ligand-free, N-acetylmannosamine-bound and substrate analog GlcNAc-bound crystal structures (2.33, 2.20, and 2.20 Å resolution, respectively). These demonstrate, in combination with small-angle X-ray scattering, that SaNanK is a dimer that adopts a closed conformation upon substrate binding. Analysis of the EXGY motif reveals that the tyrosine binds to the N-acetyl group to select for the "boat" conformation of N-acetylmannosamine. Moreover, SaNanK has a stacked arginine pair coordinated by negative residues critical for thermal stability and catalysis. These combined elements serve to constrain the active site and orient the substrate in lieu of Zn binding, representing a significant departure from canonical NanK binding. This characterization provides insight into differences in the ROK family and highlights a novel area for antimicrobial discovery to fight Gram-positive and S. aureus infections.

Bibliographical note

© 2020 Coombes et al.

Discover related content

Find related publications, people, projects, datasets and more using interactive charts.

View graph of relations