A Global Approach to Identify Novel Broad-Spectrum Antibacterial Targets among Proteins of Unknown Function

Abstract

Attempted allelic replacement of 144 <i>Streptococcus pneumoniae</i> open reading frames of previously uncharacterized function led to the identification of 36 genes essential for growth under laboratory conditions. Of these, 14 genes <i>(obg</i>, <i>spoIIIJ2</i>, <i>trmU</i>, <i>yacA</i>, <i>yacM</i>, <i>ydiC</i>, <i>ydiE</i>, <i>yjbN</i>, <i>yneS</i>, <i>yphC</i>, <i>ysxC</i>, <i>ytaG</i>, <i>yloI</i> and <i>yxeH4)</i> were also essential in <i>Staphylococcus aureus</i> and <i>Haemophilus influenzae </i>or<i> Escherichia coli,</i> 2 genes <i>(yrrK</i> and <i>ydiB)</i> were only essential in <i>H. influenzae</i> as well as <i>S. pneumoniae</i> and 8 genes were necessary for growth of <i>S.</i><i>pneumoniae</i> and <i>S. aureus </i>and did not have a homolog in <i>H. influenzae</i><i>(murD2</i>, <i>ykqC</i>, <i>ylqF</i>, <i>yqeH</i>, <i>ytgP</i>, <i>yybQ)</i> or were not essential in that organism <i>(yqeL, yhcT)</i>. The proteins encoded by these genes could represent good targets for novel antibiotics covering different therapeutic profiles. The putative functions of some of these essential proteins, inferred by bioinformatic analysis, are presented. Four mutants, with deletions of loci not essential for in vitro growth, were found to be severely attenuated in a murine respiratory tract infection model, suggesting that not all targets for antibacterial therapeutics are revealed by simple in vitro essentiality testing. The results of our experiments together with those collated from previously reported studies including <i>Bacillus subtilis</i>, <i>E. coli</i> and <i>Mycoplasma </i>sp<i>.</i> demonstrate that gene conservation amongst bacteria does not necessarily indicate that essentiality in one organism can be extrapolated to others. Moreover, this study demonstrates that different experimental procedures can produce apparently contradictory results.