Date of Degree
Biochemistry, Biophysics, and Structural Biology
β-lactamases are enzymes produced by bacteria resistant to antibiotics. A common feature on beta lactam antibiotics is the beta-lactam ring. β-lactamases hydrolyze the β-lactam ring leaving the antibiotic inoperative. The advent of bacteria that are resistant to β-lactams has impelled researchers to find inhibitors for β-lactamases that mimic the lactam ring but do not get hydrolyzed. One group of these new antibiotics is the aryl boronic acids. The main reason the boronic acids have been chosen as potential drugs is their lack of toxicity and their easy excretion in the urine. One of the most important structural features of these compounds is their chemical and geometric fitness in the active site of β-lactamases. Boronic acids mimic the tetrahedral intermediate formed in the half-acylation reaction that occurs during the hydrolysis of the β-lactam ring. The major goal of the research presented here was to discover new aryl boronic acids inhibitors of penicillinases from the class A β-lactamases. To accomplish this goal, commercially available boronic acids that are manufactured for the Suzuki reaction were used. These compounds included fluorinated, chlorinated, brominated, carboxylated, nitrophenylated, pinacol-esterified and thiophene-carboxylated aryl boronic acid derivatives. Kinetic evaluations of each class of compounds were performed under pseudo first-order enzymatic reaction conditions and the inhibitory constants (Ki) were reported using nitrocefin as substrate for two enzymes: the in-house expressed β-lactamase BlaC and the β-lactamase from Bacillus cereus 569/H9 (Calbiochem) identified as TEM-116. The structure-activity relationship (SAR) showed that the most potent inhibitors of BlaC β-lactamase were 2-carboxythiophene-5-boronic acid; 3,4,5-trifluorophenylboronic acid; 3-nitrophenytlboronic acid and 2,3,4,5- tetrafluorophenylboronic acid, Ki values of 1.2, 175.7, 213.9 and 228.6 micromolar respectively. In addition, SAR revealed that the most potent inhibitors for Bacillus cereus β-lactamase I were 2-carboxythiophen-5-boronic acid, 3-carboxyphenylboronic acid, 2-carboxythiophene-4-boronic acid, and 3-carboxy-4-fluorophenylboronic acid having Ki values of 1.1, 19.4, 46.5, and 47.1 micromolar respectively. To gain further insight into the molecular interactions between each class of inhibitors and their targeted enzymes docking experiments were performed using Autodock Vina program combined with Sculpt from MDL and followed by the molecular visualization of the protein-ligand complexes using Swiss-PdbViewer and DiscoveryStudio from Accelerys. The results conclusively show that some selective classes of aryl boronic acids are potent competitive inhibitors of BlaC and Bacillus cereus β-lactamase I and that they should be further considered for advanced drug discovery and improvement of treatment against antibiotic resistant bacteria. Furthermore, the discovery that 4,4’-DDT is an inhibitor of Mycobacterium tuberculosis β-lactamase, combined with in silico studies, suggests that further elaboration of this molecule may be one route to new inhibitors.
Barquero, Juan F., "Boronic Acids as Penicillinase Inhibitors" (2013). CUNY Academic Works.