A pyrophosphate bridge links the pyruvate-containing secondary cell wall polymer of Paenibacillus alvei CCM 2051 to muramic acid

The peptidoglycan, the secondary cell wall polymer (SCWP), and the surface layer (S-layer) glycoprotein are the major glycosylated cell wall component s of Paenibacillus alvei CCM 2051. In this report, the complete structure o f the SCWP, its linkage to the peptidoglycan layer, and its physicochemical properties have been investigated. From the combined evidence of chemical and structural analyses together with one- and two-dimensional nuclear magn etic resonance spectroscopy, the following structure of the SCWP-peptidogly can complex is proposed: [(Pyr4,6)-beta -D-ManpNAc-(1 -->4)-beta -D-GlcpNAc-(1 -->3)](n similar to 1 1)-(Pyr4,6)-beta -D-ManpNAc-(1 -->4)-alpha -D-GlcpNAc-(1 -->O)-PO2-O-PO2-(O -->6)-MurNAc- Each disaccharide unit is substituted by 4,6-linked pyruvic acid residues. Under mild acidic conditions, up to 50% of them are lost, leaving non-subst ituted ManNAc residues. The anionic glycan chains constituting the SCWP are randomly linked via pyrophosphate groups to C-6 of muramic acid residues o f the peptidoglycan layer. P-31 NMR reveals two signals that, as a conseque nce of micelle formation, experience different line broadening. Therefore, their integral ratio deviates significantly from 1:1. By treatment with eth ylenediaminetetraacetic acid, sodium dodecyl sulfate, and sonication immedi ately prior to NMR measurement, this ratio approaches unity. The reversibil ity of this behavior corroborates the presence of a pyrophosphate linker in this SCWP-peptidoglycan complex. In addition to the determination of the structure and linkage of the SCWP, a possible scenario for its biological function is discussed.