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Volume 162, Issue 6

Conserved transmembrane glycine residues in the polysaccharide co-polymerase protein WzzB influence protein–protein interactions Free

Abstract

The O antigen (Oag) component of lipopolysaccharides (LPS) is crucial for virulence and Oag chain-length regulation is controlled by the polysaccharide co-polymerase class 1 (PCP1) proteins. Crystal structure analyses indicate that structural conservation among PCP1 proteins is highly maintained, however the mechanism of Oag modal-chain-length control remains to be fully elucidated. PCP1 protein WzzB confers a modal-chain length of 10–17 Oag repeat units (RUs), whereas the PCP1 protein WzzB confers a modal-chain length of ~16–28 Oag RUs. Both proteins share >70 % overall sequence identity and contain two transmembrane (TM1 and TM2) regions, whereby a conserved proline-glycine-rich motif overlapping the TM2 region is identical in both proteins. Conserved glycine residues within TM2 are functionally important, as glycine to alanine substitutions at positions 305 and 311 confer very short Oag modal-chain length (~2–6 Oag RUs). In this study, WzzB was co-expressed with WzzB in and a single intermediate modal-chain length of ~11–21 Oag RUs was observed, suggesting the presence of Wzz:Wzz interactions. Interestingly, co-expression of WzzB with WzzB conferred a bimodal LPS Oag chain length (despite over 99 % protein sequence identity), and we hypothesized that the proteins fail to interact. Co-purification assays detected His-WzzB co-purifying with FLAG-tagged WzzB but not with FLAG-tagged WzzB, supporting our hypothesis. These data indicate that the conserved glycine residues in TM2 are involved in Wzz:Wzz interactions, and provide insight into key interactions that drive Oag modal length control.

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Keyword(s): Lipopolysaccharide , O antigen , polysaccharide co-polymerase , Shigella flexneri , transmembrane glycines and Wzz proteins
© 2016 The Authors
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2016-06-01
2024-04-20
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Conserved transmembrane glycine residues in the Shigella flexneri polysaccharide co-polymerase protein WzzB influence protein–protein interactions
Microbiology 162, 921 (2016); https://doi.org/10.1099/mic.0.000282
/content/journal/micro/10.1099/mic.0.000282
/content/journal/micro/10.1099/mic.0.000282
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