Screening of β-Lactamase Inhibitors Using Phytochemicals with recombinant β-Lactamase

Abstract The emergence and rapid spread of β-lactamase-mediated antibiotic resistance poses a significant challenge to the effectiveness of β-lactam antibiotics, highlighting the need for exploring β-lactamase inhibitory agents. The present study aimed to develop a recombinant TEM β-lactamase and evaluate natural plant extracts for their β-lactamase inhibitory potential. A conserved blaTEM sequence (DQ849322.1) was codon-optimized, cloned into the pET-28a (+) expression vector and over expressed in Escherichia coli BL21 (DE3). A specific activity of 211 IU/mg was observed for the purified recombinant enzyme. The purified enzyme was used for evaluating the inhibitory potential of 60 herbal and spices aqueous extracts through nitrocefin assay and microbial inhibition assay. Extracts from Custard apple ( Annona squamosa) seeds, Java Plum ( Syzygium cumini) fruit, Dry lemon zest ( Citrus aurantifolia) , Clove ( Syzygium aromaticum) , Winter cherry ( Withania somnifera) roots, Cinchona leaves ( Cinchona pubescens) , and Carom seeds ( Trachyspermum ammi) exhibited higher β-lactamase inhibition. The selected extracts had an inhibition percentage of 78.60 ± 0.19%, 77.89 ± 0.16%, 77.80 ± 0.20%, 75.31 ± 0.15%, 71.31 ± 0.30%, 67.32 ± 0.17% and 66.87 ± 0.11%, respectively (p < 0.0001). Amoxicillin resistant E. coli , improved sensitivity, when tested along with inhibitors. The results suggest that plant-derived bioactive phytochemical constituents possess potential β-lactamase inhibitory activity and may serve as valuable sources for antibiotic resistance-modifying agents.