The gene bglH present in the bgl operon of Escherichia coli, responsible for uptake and fermentation of beta-glucosides encodes for a carbohydrate-specific outer membrane porin

Mol Microbiol. 1999 Jan;31(2):499-510. doi: 10.1046/j.1365-2958.1999.01191.x.

Abstract

The cryptic gene bglH from the Escherichia coli chromosome was cloned into a tacOP-driven expression vector. The resulting plasmid was transferred into the porin-deficient E. coli strain KS26 and the protein was expressed by addition of IPTG. The BglH protein was localized in the outer membrane. It was purified to homogeneity using standard methods. Reconstitution experiments with lipid bilayer membranes defined BglH as a channel-forming component, i.e. it is an outer membrane porin. The single-channel conductance of BglH (560 pS in 1 M KCl) was only one-third of that of the general diffusion porins of E. coli outer membrane. The presence of carbohydrates in the aqueous phase led to a dose-dependent block of ion transport through the channel, similar to that found for LamB (maltoporin) of E. coli and Salmonella typhimurium, which means that BglH is a porin specific for the uptake of carbohydrates. The binding constants of a variety of different carbohydrates were calculated from titration experiments of the BglH-induced membrane conductance. The tightest binding was observed with the aromatic beta-D-glucosides arbutin and salicin, and with gentibiose and cellobiose. Binding of maltooligosaccharides to BglH was in contrast to their binding to LamB in that it was much weaker, indicating that the binding site of BglH for carbohydrates is different from that of LamB (maltoporin). The kinetics of cellopentaose binding to BglH was investigated using the carbohydrate-induced current noise and was compared with that of cellopentaose binding to LamB (maltoporin) and ScrY (sucroseporin).

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Carbohydrate Metabolism*
  • Escherichia coli / genetics
  • Escherichia coli / metabolism*
  • Fermentation
  • Genes, Bacterial
  • Glucosidases / chemistry
  • Glucosidases / genetics
  • Glucosidases / isolation & purification
  • Glucosidases / metabolism*
  • Glucosides / metabolism*
  • Lipid Bilayers / metabolism
  • Molecular Sequence Data
  • Oligosaccharides / metabolism
  • Operon*
  • Porins / metabolism*

Substances

  • Glucosides
  • Lipid Bilayers
  • Oligosaccharides
  • Porins
  • maltopentaose
  • Glucosidases
  • 6-phospho-beta-glucosidase