The Haemophilus influenzae HMW1 adhesin is glycosylated in a process that requires HMW1C and phosphoglucomutase, an enzyme involved in lipooligosaccharide biosynthesis

Mol Microbiol. 2003 May;48(3):737-51. doi: 10.1046/j.1365-2958.2003.03450.x.

Abstract

Non-typeable Haemophilus influenzae is a common respiratory pathogen and an important cause of morbidity in humans. The non-typeable H. influenzae HMW1 and HMW2 adhesins are related proteins that mediate attachment to human epithelial cells, an essential step in the pathogenesis of disease. Secretion of these adhesins requires accessory proteins called HMW1B/HMW2B and HMW1C/HMW2C. In the present study, we investigated the specific function of HMW1C. Examination of mutant constructs demonstrated that HMW1C influences both the size and the secretion of HMW1. Co-immunoprecipitation and yeast two-hybrid assays revealed that HMW1C interacts with HMW1 and forms a complex in the cytoplasm. Additional experiments and homology analysis established that HMW1C is required for glycosylation of HMW1 and may have glycotransferase activity. The glycan structure contains galactose, glucose and mannose and appears to be generated in part by phosphoglucomutase, an enzyme important for lipooligosaccharide biosynthesis. In the absence of glycosylation, HMW1 is partially degraded and is efficiently released from the surface of the organism, resulting in reduced adherence. Based on these results, we conclude that glycosylation is a prerequisite for HMW1 stability. In addition, glycosylation appears to be essential for optimal HMW1 tethering to the bacterial surface, which in turn is required for HMW1-mediated adherence, thus revealing a novel mechanism by which glycosylation influences cell-cell interactions.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Adhesins, Bacterial / metabolism*
  • Animals
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism*
  • Cell Adhesion / physiology
  • Cell Line
  • Epithelial Cells / metabolism
  • Epithelial Cells / microbiology
  • Glycosylation
  • Haemophilus influenzae / chemistry
  • Haemophilus influenzae / metabolism*
  • Humans
  • Lipopolysaccharides / biosynthesis*
  • Mutation
  • Phosphoglucomutase / metabolism*
  • Recombinant Fusion Proteins / metabolism
  • Two-Hybrid System Techniques

Substances

  • Adhesins, Bacterial
  • Bacterial Proteins
  • HMW1 protein, bacteria
  • Lipopolysaccharides
  • Recombinant Fusion Proteins
  • lipid-linked oligosaccharides
  • Phosphoglucomutase