Author(s): MULINARI, F.; BECKER-RITT, A. B.; DEMARTINI, D. R.; LIGABUE-BRAUN, R.; STANISÇUASKI, F.; VERLI, H.; FRAGOSO, R. R.; SCHROEDER, E. K.; CARLINI, C. R.; GROSSI-de-SÁ, M. F.

Summary: Ureases, nickel-dependent enzymes that catalyze the hydrolysis of urea into ammonia and bicarbonate, are widespread in plants, bacteria, and fungi. Previously, we cloned a cDNA encoding a Canavalia ensiformis urease isoform named JBURE-II, corresponding to a putative smaller urease protein (78 kDa) when compared to other plant ureases. Aiming to produce the recombinant protein, we obtained jbure-IIb, with different 3? and 5? ends, encoding a 90 kDa urease. Three peptides unique to the JBURE-II/-IIb protein were detected by mass spectrometry in seed extracts, indicating that jbure-II/-IIb is a functional gene. Comparative modeling indicates that JBURE-IIb urease has an overall shape almost identical to C. ensiformis major urease JBURE-I with all residues critical for urease activity. The cDNA was cloned into the pET101 vector and the recombinant protein was produced in Escherichia coli. The JBURE-IIb protein, although enzymatically inactive presumably due to the absence of Ni atoms in its active site, impaired the growth of a phytopathogenic fungus and showed entomotoxic properties, inhibiting diuresis of Rhodnius prolixus isolated Malpighian tubules, in concentrations similar to those reported for JBURE-I and canatoxin. The antifungal and entomotoxic properties of the recombinant JBURE-IIb apourease are consistent with a protective role of ureases in plants.

Publication year: 2011

Types of publication: Journal article

Unit: Embrapa Genetic Resources & Biotechnology

Keywords: Canavalia Ensiformis