The development of new antimicrobial agents is failing to keep pace with the rapid spread of bacterial resistance, which represents an increasing global health burden. Topoisomerases, including the bacterial enzymes DNA gyrase and topoisomerase IV, regulate the topological state of DNA molecules and play a key role in replication and transcription processes. An increasing number of bacteria have also developed resistance to fluoroquinolones, inhibitors of DNA gyrase and topoisomerase IV, which are among the most prescribed antibacterial agents. Due to the emergence of resistance, research has shifted toward the development of new inhibitors, including allosteric inhibitors, which are the focus of this master’s thesis. Allosteric inhibitors of DNA gyrase bind to a new hydrophobic pocket at the interface between the GyrA and GyrB subunits, which differs from the fluoroquinolone binding site. Therefore, they represent a promising approach for inhibiting bacterial strains that have already developed resistance to fluoroquinolones. In contrast to fluoroquinolones, which promote DNA cleavage, allosteric inhibitors suppress this activity.
As part of this master’s thesis, molecular docking studies were performed on analogues of Gyr04, a recently discovered thiazole-based DNA gyrase inhibitor developed at the Faculty of Pharmacy, University of Ljubljana, targeting the allosteric binding site. The results showed that the most promising analogues were those with a halogen, methyl, or methoxy group attached to the aniline ring, and a methoxyphenyl or formylphenyl substituent attached to the thiazole ring. These findings guided further experimental work, in which eight new thiazole-based allosteric inhibitors of DNA gyrase were synthesised, analysed, and biologically evaluated.
The results of the inhibitory activity measurements against DNA gyrase from Escherichia coli showed that two compounds exhibited inhibitory activity. Upon the addition of compound 5a (10 μM) 21% of the enzymatic activity remained, while compound 5g (10 μm) reduced the activity to only 13%. Both compounds have a bromine atom attached to the thiazole ring; compound 5a has two methyl groups on the aniline ring, whereas compound 5g has a chlorine atom. The other inhibitors exhibited weaker inhibitory effects, as the residual enzymatic activity remained above 50% after the addition of 10 μM of the compound. Within this master’s thesis, preliminary studies of Gyr04 analogues were conducted. The results provide a basis for future research in this field and contribute to the development of new antibacterial agents aimed at combating bacterial resistance.
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