Cross-Link Formation and Peptidoglycan Lattice Assembly in the FemA Mutant of Staphylococcus aureus

[Image: see text] Staphylococcus aureus FemA mutant grown in the presence of an alanine-racemase inhibitor was labeled with d-[1-(13)C]alanine, l-[3-(13)C]alanine, [2-(13)C]glycine, and l-[5-(19)F]lysine to characterize some details of the peptidoglycan tertiary structure. Rotational-echo double-resonance (REDOR) NMR of isolated cell walls was used to measure internuclear distances between (13)C-labeled alanines and (19)F-labeled lysine incorporated in the peptidoglycan. The alanyl (13)C labels were preselected for REDOR measurement by their proximity to the glycine label using (13)C–(13)C spin diffusion. The observed (13)C–(13)C and (13)C–(19)F distances are consistent with a tightly packed, hybrid architecture containing both parallel and perpendicular stems in a repeating structural motif within the peptidoglycan.