UVI31+: A novel UV-inducible protein in Chlamydomonas reinhardtii showing β-lactamase activity

Renu Minda, Devika Sirohi, Jacinta S. D'Souza, S.K. Bhattacharjee and B.J. Rao

Department of Biological Sciences, Tata Institute of Fundamental Research, Homi Bhabha Road, Colaba, Mumbai-400005, INDIA

Chlamydomonas reinhardtii exhibits classical programmed cell death response following UV-stress. The dying cells seem to impart protection against impending UV-stress on normal unexposed cells, thereby perhaps revealing that apoptosis is an adaptive response to UV stress. In the pursuit of understanding cellular responses to UV stress, we cloned and expressed one of the UV-inducible genes (uvi31+). The protein expressed in E.coli from uvi31+ cDNA was purified and characterized. Computational analysis showed that the protein contained a Bol-A like domain. BolA from Escherichia coli has been well characterized and confers round morphology when overexpressed. Most surprisingly, UVI31+ from C. reinhardtii demonstrated penicillin inactivating property. The beta-lactamase activity was indeed associated with uvi31+ gene as evidenced by the biological assays specifically with uvi31+ gene harboring plasmids. Purified protein activity was primarily against penicillins, with no detectable activity against the tested carbapenems, monobactams, and cephalosporins. Microiodometric assay confirmed the formation of ampicilloic acid on hydrolysis of the β-lactam antibiotic by UVI31+ action. Biochemical analysis with penicillins showed that its kcat/Km values ranged between 6.1 and 6.9 x 102 uM-1 s-1. To the best of our knowledge this is the first report showing that. We now believe that the protein, a BolA domain containing β-lactamase may indeed carry other functions as well that relate to UV-stress response pathways and we propose a model that highlights its central role in C. reinhardti.

e-mail address of presenting author: bjrao@tifr.res.in