posted on 2018-05-28, 00:00authored byJoseph
A. H. Romaniuk, Lynette Cegelski
Gram-positive
bacteria surround themselves with a multilayered
macromolecular cell wall that is essential to cell survival and serves
as a major target for antibiotics. The cell wall of Staphylococcus
aureus is composed of two major structural components, peptidoglycan
(PG) and wall teichoic acid (WTA), together creating a heterogeneous
and insoluble matrix that poses a challenge to quantitative compositional
analysis. Here, we present 13C cross polarization magic
angle spinning solid-state nuclear magnetic resonance (NMR) spectra
of intact cell walls, purified PG, and purified WTA. The spectra reveal
the clear molecular differences in the two polymers and enable quantification
of PG and WTA in isolated cell walls, an attractive alternative to
estimating teichoic acid content from a phosphate analysis of completely
pyrolyzed cell walls. Furthermore, we discovered that unique PG and
WTA spectral signatures could be identified in whole-cell NMR spectra
and used to compare PG and WTA levels among intact bacterial cell
samples. The distinguishing whole-cell 13C NMR contributions
associated with PG include the GlcNAc-MurNAc sugar carbons and glycyl
α-carbons. WTA contributes carbons from the phosphoribitol backbone.
Distinguishing 15N spectral signatures include glycyl amide
nitrogens in PG and the esterified d-alanyl amine nitrogens
in WTA. 13C NMR analysis was performed with samples at
natural abundance and included 10 whole-cell sample comparisons. Changes
consistent with altered PG and WTA content were detected in whole-cell
spectra of bacteria harvested at different growth times and in cells
treated with tunicamycin. This use of whole-cell NMR provides quantitative
parameters of composition in the context of whole-cell activity.