Abstract
EGS (external guide sequence) technology is a promising approach to designing new antibiotics. EGSs are short antisense oligoribonucleotides that induce RNase P-mediated cleavage of a target RNA by forming a precursor tRNA-like complex. The ftsZ mRNA secondary structure was modeled and EGSs complementary to two regions with high probability of being suitable targets were designed. In vitro reactions showed that EGSs targeting these regions bound ftsZ mRNA and elicited RNase P-mediated cleavage of ftsZ mRNA. A recombinant plasmid, pEGSb1, coding for an EGS that targets region “b” under the control of the T7 promoter was generated. Upon introduction of this plasmid into Escherichia coli BL21(DE3)(pLysS) the transformant strain formed filaments when expression of the EGS was induced. Concomitantly, E. coli harboring pEGSb1 showed a modest but significant inhibition of growth when synthesis of the EGSb1 was induced. Our results indicate that EGS technology could be a viable strategy to generate new antimicrobials targeting ftsZ.
Highlights
Bacterial cell division is a complex process that occurs following the replication and segregation of chromosomal DNA to the two halves of the growing cell
The E. coli ftsZ gene is located towards the distal end of the dcw cluster, a group of 16 genes involved in cell division and cell wall synthesis [37]
The quick increase in resistant strains observed among a number of bacterial pathogens together with the low number of candidate compounds existing in the pipeline warrant the need to look for alternatives to design new antibiotics [45,46]
Summary
Bacterial cell division is a complex process that occurs following the replication and segregation of chromosomal DNA to the two halves of the growing cell. In the case of Gram negative bacteria, the division process requires at least 14 cytoplasmic, membrane and periplasmic proteins, of which 10 are essential [1,2,3,4,5] These proteins form a structure known as the divisome, a ring-like cell division complex located at midcell that constricts during division and disappears when the cells separate [1,2,3,4,5]. The most conserved of all known bacterial cell division genes is the proto-ring protein FtsZ, which functions as scaffold for the divisome and generates the constrictive force to initiate division of the cell [6,9,10]. Numerous reports have been published proposing novel cell division inhibitors that act by blocking FtsZ and hold high therapeutic potential but none of them have been fully developed and released to the market to date [11,12,13,14,15,16,17,18]
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