Characterization of a promiscuous glutamate cysteine ligase from Leptospira interrogans

SASONI N; GUERRERO SA; IGLESIAS AA; ARIAS DG

Rosario

Congreso; Reunión de la SAIB (50 reunión); 2014

Glutathione (GSH) is the most abundant low molecular weight thiol in almost all eukaryotic cells as well as in proteo and cyanobacteria. It is synthesized enzymatically in two ATP?Mg+2 dependent steps. First, glutamate cysteine ligase (GCL) establishes a peptide bond between cysteine and glutamate, forming γ-glutamylcysteine (Υ?GC). Second, glutathione synthetase (GS) adds a glycine residue to the carboxy-terminus of Υ?GC, producing glutathione. Analysis of the genome sequences of Leptospira interrongans (the casusative agent of the leptospirosis) indicates the absence of the encoding-gene to GS. However, the presence of the gene that encodes GCL (LIC11812) leads us to believe that L. interrogans could has Υ?GC as redox buffer similar to Halobacteria and Lactic acid bacteria. Therefore, we cloned, expressed in recombinant form and characterized the functionality of GCL of L.interrogans. We showed that the enzyme is active in presence of their physiologic substrates (glutamate, cysteine and ATP), and also has the ability to use GTP, aspartic and serine, although in lower proportion. The enzyme has optimal activity atpH~7.5. Our results add value to the genome project information and contribute to the understanding of the redox metabolism present in this bacterium.