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    Cloning and functional expression of secreted phospholipases A2 from Bothrops diporus (Yarará Chica)

    Year: 2012

    Journal: Biochemical and Biophysical Research Communications, 2012, 427 (2), 321-325, 20131009

    Authors: Pablo Javier Yunes Quartino, José Luis Barra, Gerardo Daniel Fidelio

    Organizations: Centro de Investigaciones en Química Biológica de Córdoba (CIQUIBIC, UNC–CONICET), Departamento de Química Biológica, Facultad de Ciencias Químicas, Universidad Nacional de Córdoba. Haya de la Torre y Medina Allende, Ciudad Universitaria, X5000HUA Córdoba, Argentina

    Bothrops diporus is a very common viper in Argentina. At present, no complete sequence of secreted phospholipase A2 (sPLA2) from this snake has been reported. We have cloned two sPLA2 isoenzymes as well as a putative sPLA2-like myotoxin from venom gland. The two sPLA2 were expressed as inclusion bodies in Escherichia coli with an N-terminal tag of ubiquitin. After in vitro renaturation and cleavage step, using an ubiquitin specific peptidase, the recombinants exhibited sPLA2 activity when analyzed by means of Langmuir dilauroylphosphatidylcholine monolayers as substrate. Both enzymes have a similar surface pressure-activity profile when compared with non-recombinant purified isoforms. To our knowledge, this is the first time that analysis of optimal lateral pressure of substrate monolayers by using the surface barostat technique is performed on recombinant sPLA2s.