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Structural and functional characterization of NanU, a novel high-affinity sialic acid-inducible binding protein of oral and gut-dwelling Bacteroidetes species

DOI: 10.1042/BJ20131415 DOI Help
PMID: 24351045 PMID Help

Authors: Chatchawal Phansopa (University of Sheffield) , Sumita Roy (University of Sheffield) , J B Rafferty (University of Sheffield) , C. W. ian Douglas (University of Sheffield) , Jagroop Pandhal (University of Sheffield) , Phillip C. Wright (University of Sheffield) , David J Kelly (University of Sheffield) , Graham p. Stafford (University of Sheffield)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Biochemical Journal , VOL 458 , PAGES 499 - 511

State: Published (Approved)
Published: March 2014
Diamond Proposal Number(s): 8987 , 7423

Open Access Open Access

Abstract: Many human-dwelling bacteria acquire sialic acid for growth or surface display. We identified previously a sialic acid utilization operon in Tannerella forsythia that includes a novel outer membrane sialic acid-transport system (NanOU), WHERE NanO (neuraminate outer membrane permease) is a putative TonB-dependent receptor and NanU (extracellular neuraminate uptake protein) is a predicted SusD family protein. Using heterologous complementation of nanOU genes into an Escherichia coli strain devoid of outer membrane sialic acid permeases, we show that the nanOU system from the gut bacterium Bacteroides fragilis is functional and demonstrate its dependence on TonB for function. We also show that nanU is required for maximal function of the transport system and that it is expressed in a sialic acid-responsive manner. We also show its cellular localization to the outer membrane using fractionation and immunofluorescence experiments. Ligand-binding studies revealed high-affinity binding of sialic acid to NanU (Kd ~400 nM) from two Bacteroidetes species as well as binding of a range of sialic acid analogues. Determination of the crystal structure of NanU revealed a monomeric SusD-like structure containing a novel motif characterized by an extended kinked helix that might determine sugar-binding specificity. The results of the present study characterize the first bacterial extracellular sialic acid-binding protein and define a sialic acid-specific PUL (polysaccharide utilization locus).

Journal Keywords: Bacterial; Bacteroides; Bacteroidetes; Biological; Crystallography; X-Ray; Escherichia; Intestines; Membrane; Mouth; Mutation; N-Acetylneuraminic; Protein; Secondary

Subject Areas: Biology and Bio-materials


Instruments: I04-1-Macromolecular Crystallography (fixed wavelength)

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