The molecular basis of regulation of bacterial capsule assembly by Wzc

Abstract

Bacterial extracellular polysaccharides (EPSs) play critical roles in virulence. Many bacteria assemble EPSs via a multi-protein “Wzx-Wzy” system, involving glycan polymerization at the outer face of the cytoplasmic/inner membrane. Gram-negative species couple polymerization with translocation across the periplasm and outer membrane and the master regulator of the system is the tyrosine autokinase, Wzc. This near atomic cryo-EM structure of dephosphorylated Wzc from E. coli shows an octameric assembly with a large central cavity formed by transmembrane helices. The tyrosine autokinase domain forms the cytoplasm region, while the periplasmic region contains small folded motifs and helical bundles. The helical bundles are essential for function, most likely through interaction with the outer membrane translocon, Wza. Autophosphorylation of the tyrosine-rich C-terminus of Wzc results in disassembly of the octamer into multiply phosphorylated monomers. We propose that the cycling between phosphorylated monomer and dephosphorylated octamer regulates glycan polymerization and translocation.

A Conserved Structural Role for the Walker-A Lysine in P-Loop Containing Kinases

“…We have previously demonstrated the unique conformational dynamics of the catalytic core of the archetypal BY-kinase, Escherichia coli (K12) Wzc (Wzc CDΔC ); these dynamics appear to have significant functional consequences. Our studies revealed the presence of two major conformations with distinct long-and short-range structural features (Hajredini et al, 2020) 2+ , is stabilized upon formation of a closed octameric ring seen both in the structure of the isolated CD (Bechet et al, 2010) as well as in the membrane-anchored full-length protein (Yang et al, 2021). Oligomer formation is necessary to enable intermolecular autophosphorylation.…”

Bacterial outer-membrane polysaccharide export (OPX) proteins occupy three structural classes with selective β-barrel porin requirements for polymer secretion

“…as per a previous report (Sande et al, 2019). have been provided as per the PDB files 2J58 (Dong et al, 2006) and 7NII (Yang et al, 2021) (respectively) for size references. Structure models for WzaB (Fig.…”

“…All proteins, spaces, and distances have been depicted at the same relative scale across a representative 327 Å periplasmic space in a M. xanthus cell. X-ray crystal structures for Wza Ec (chain A) and Wzc Ec (chains F and G) have been provided as per the PDB files 2J58 (Dong et al ., 2006) and 7NII (Yang et al ., 2021) (respectively) for size references. Structure models for WzaB (Fig.…”

“…The Wzb bacterial tyrosine phosphatase (BYP) protein in turn regulates the state of PCP-2A Wzc and PCP-2B- associated Wze phosphorylation (Mori et al ., 2012). Wzb-mediated dephosphorylation of BYK domains has been proposed to drive Wzc octamerization, in turn affecting Wzy-mediated polymerization and interaction with the outer-membrane polysaccharide export (OPX) Wza translocon needed for polymer transport through the periplasm and across the outer membrane (OM) (Yang et al ., 2021) ( Fig. 1A ).…”

Tyrosine Kinases and Phosphatases: Enablers of the Porphyromonas gingivalis Lifestyle

“…Wzc autophosphorylation causes octamer dissociation leading to closure of the Wza translocon. Reactivation then depends on the dephosphorylation of Wzc by Wzb [ 21 ]. Detailed mechanistic studies have not been performed in P. gingivalis ; however, deletion of the ptk1 gene results in loss of EPS secretion in strain 33277, and failure to produce a capsule in strain W83 [ 15 ].…”