HMW1C-like proteins in two categories: Those encoded by loci that contain obvious substrate genes and those encoded by isolated genes without adjacent substrate genes.

<p>Numbers below <i>hmw1C</i>-like genes represent translated protein sequence percent identity/similarity when compared to <i>H. influenzae</i> HMW1C. (<b>A</b>) HMW1C-like enzymes encoded in apparent TPS systems. (<b>B</b>) HMW1C-like enzymes encoded in loci without obvious surface protein targets for glycosylation. Abbreviations: <i>Hi</i>, <i>H. influenzae</i> 86-028NP; <i>Bsp</i>, <i>Burkholderia</i> species GCE1003; <i>Ec</i>, Enterotoxigenic <i>E. coli</i> H10407; <i>Yp</i>, <i>Y. pseudotuberculosis</i> YPIII; <i>Ap</i>, <i>Actinobacillus pleuropneumoniae</i> L20; <i>Hd</i>, <i>H. ducreyi</i> HD35000; <i>Kk</i>, <i>K. kingae</i> 269–492; <i>hyp</i>, hypothetical with no conserved domains; <i>hyp</i>¹, predicted lipoprotein; <i>hyp</i>², predicted UDP-glcNAc carboxyvinyltransferase; <i>hyp</i>³, predicted 2 C-methyl-D erythritol-4-phosphate cytidyltransferase; <i>hyp</i>⁴, predicted deoxyguanosinetriphosphate triphosphohydrolase.</p

Glycosylation by HMW1C may play several roles in promoting HMW1 stability, export, folding, and function.

<p>HMW1C has the potential to contribute to several different processes that occur during HMW1 synthesis and transit across the inner and outer membranes. First of all, HMW1C glycosylates the HMW1 adhesin in the cytoplasm and is likely to be involved in the stability of the HMW1 adhesin during or after its synthesis. Glycosylation may contribute to stability of HMW1 in the (<b>A</b>) cytoplasm or (<b>C</b>) periplasm <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1003977#ppat.1003977-Grass1" target="_blank">[5]</a>, <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1003977#ppat.1003977-StGeme1" target="_blank">[7]</a>. Alternatively, the HMW1C protein may improve stability of HMW1 by acting as a (<b>B</b>) chaperone prior to secretion of the adhesin. It is unlikely that the activity of HMW1C is required for export of the adhesin across either the inner or outer membrane, as fully processed HMW1 is found in the supernatant in the absence of HMW1C <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1003977#ppat.1003977-Grass1" target="_blank">[5]</a>. It is unclear whether glycosylation influences interaction of HMW1 with the (<b>D</b>) HMW1B periplasmic domain prior to transit, (<b>E</b>) the HMW1B pore during transit, or (<b>F</b>) the docking region of HMW1B upon surface tethering <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1003977#ppat.1003977-Grass1" target="_blank">[5]</a>, <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1003977#ppat.1003977-Buscher2" target="_blank">[11]</a>. It is also unclear whether glycosylation participates in (<b>G</b>) protein folding upon export. Evidence from the nonglycosylated <i>Bordetella</i> prototypic, two-partner, secreted adhesin FHA indicates that this adhesin remains unfolded in the cytoplasm and folds very rapidly upon export via its TpsB secretion pore <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1003977#ppat.1003977-Hodak1" target="_blank">[30]</a>. One hypothesis is that the energy generated by this rapid folding is at least part of what drives export of TpsA proteins across the outer membrane <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1003977#ppat.1003977-JacobDubuisson1" target="_blank">[2]</a>. Finally, glycosylation of HMW1 may be required for (<b>H</b>) adherence to host cells or host interaction in a particular niche.</p