β-Lactamases (EC 3.5.2.6) confer resistance against β-lactam group-containing antibiotics in bacteria and higher eukaryotes, including humans. Pathogenic bacterial resistance against β-lactam antibiotics is a primary concern for potential therapeutic developments and drug targets. Here, we report putative β-lactamase activity, sulbactam binding (a β-lactam analogue) in the low μM affinity range, and site-specific interaction studies of a 14 kDa UV- and dark-inducible protein (abbreviated as UVI31+, a BolA homologue) from Chlamydomonas reinhartii. Intriguingly, the solution NMR structure of UVI31 + bears no resemblance to other known β-lactamases; however, the sulbactam binding is found at two sites rich in positively charged residues, mainly at the L2 loop regions and the N-terminus. Using NMR spectroscopy, ITC and MD simulations, we map the ligand binding sites in UVI31 + providing atomic-level insights into its β-lactamase activity. Current study is the first report on β-lactamase activity of UVI31+, a BolA analogue, from C. reinhartii. Furthermore, our mutation studies reveal that the active site serine-55 is crucial for β-lactamase activity.

Department of Bioinformatics Manipal School of Life Sciences Manipal Academy of Higher Education Manipal 576104 India

Department of Cellular Biochemistry Max Planck Institute of Biochemistry Am Klopferspitz 18 82152 Martinsried Germany

Department of Chemical Sciences Indian Institute of Science Education and Research Berhampur 760010 Odisha India

Department of Chemical Sciences Tata Institute of Fundamental Research Mumbai 400005 India

Department of Chemistry VIT Bhopal University Bhopal India

Institute of Chemistry and Metabolomics University of Luebeck 23562 Luebeck Germany

National Centre for Biomolecular Research Faculty of Science Masaryk University Kamenice 5 62500 Brno Czech Republic

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