Cloning and sequencing of the Klebsiella pneumoniae tonB gene and characterization of Escherichia coli-K. pneumoniae TonB hybrid proteins

Abstract

The tonB gene of Klebsiella pneumoniae was cloned on a 1.4-kb EcoRV fragment and expressed in Escherichia coli. Determination of the nucleotide (nt) sequence of the cloned fragment identified tonB as an open reading frame (ORF) of 729 nt. The TonB protein consists of 243 amino acids (aa) and is highly homologous to Enterobacter aerogenes TonB. The genetic context of K. pneumoniae and En. aerogenes tonB is the same, but is different from that of E. coli, Salmonella typhimurium and Serratia marcescens. K. pneumoniae tonB complemented an E. coli tonB mutant as efficiently as E. coli tonB when cloned on the same type of plasmid. Replacement of the highly hydrophobic C terminus of E. coli TonB by the weakly hydrophobic C terminus of K. pneumoniae TonB yielded a hybrid protein of the same activity as E. coli wild-type TonB. The result shows that the strong hydrophobicity of the C-terminal end is not necessary for the function of E. coli TonB.