In Gram-negative bacteria the β-barrel assembly machinery (BAM) folds and inserts proteins into the outer membrane. Without the BAM complex, β-barrels are not inserted, leading to membrane disruption and demise of the cell. Thus, an increase in the understanding of the BAM complex might help the identification of novel antimicrobial targets. The Escherichia coli BAM complex consists of 5 subunits: BamA; BamB; BamC; BamD; and BamE. Many studies have highlighted the importance of the essential components, BamA and BamD. However, the functions of the non-essential BAM subunits, BamB, BamC and BamE are unknown. In this study the transposon-directed insertion site sequencing (TraDIS) approach was used to compare insertion frequencies in mutants defective in one of the non-essential genes with the parent strain. Genes with few or no transposon insertions were suggested to be essential while genes with numerous insertions were classed as non-essential.
TraDIS is a high-throughput technique that involves sequencing of the insertion sites of a high density transposon insertion library. Libraries were produced in mutants defective in bamB, bamC or bamE. In a ∆bamB mutant, 36 genes were found to be essential that were not essential in the parent strain. The corresponding results for bamC and bamE mutants were 48 and 18, respectively. Preliminary analysis revealed that genes involved in cell division and lipopolysaccharide assembly have increased importance in a ∆bamB mutant while many genes of unknown function have increased importance in ∆bamC and ∆bamE mutants. Thus, the TraDIS approach can be used to yield insights into the biogenesis of the bacterial outer membrane. The data illustrate how the Tradis approach can be used to help identify potential targets for antibacterial drug development.