Attachment of phosphorylcholine residues to pneumococcal teichoic acids and modification of substitution patterns by the phosphorylcholine esterase.

The bacterial lung pathogen has a unique nutritional requirement for exogenous choline and attaches phosphorylcholine (-Cho) residues to the GalNAc moieties of its teichoic acids (TAs) in its cell wall. Two phosphorylcholine transferases, LicD1 and LicD2, mediate the attachment of -Cho to the O-6 positions of the two GalNAc residues present in each repeating unit of pneumococcal TAs (pnTAs), of which only LicD1 has been determined to be essential. At the molecular level, the specificity of the -Cho attachment to pnTAs by LicD1 and LicD2 remains still elusive. Here, using detailed structural analyses of pnTAs from a LicD2-deficient strain, we confirmed the specificity in the attachment of -Cho residues to pnTA. LicD1 solely transfers -Cho to α-d-GalNAc moieties, whereas LicD2 attaches -Cho to β-d-GalNAc. Further, we investigated the role of the pneumococcal phosphorylcholine esterase (Pce) in the modification of the -Cho substitution pattern of pnTAs. To clarify the specificity of Pce-mediated -Cho hydrolysis, we evaluated different concentrations and pH conditions for the treatment of pneumococcal lipoteichoic acid with purified Pce. We show that Pce can hydrolyze both -Cho residues of the terminal repeat of the pnTA chain and almost all -Cho residues bound to β-d-GalNAc However, hydrolysis was restricted to the terminal repeat. In summary, our findings indicate that LicD1 and LicD2 specifically transfer -Cho to α-d-GalNAc and β-d-GalNAc moieties, respectively, and that Pce removes distinct -Cho substituents from pnTAs.