FIRST CHARACTERIZATION OF A NITRIC OXIDE SYNTHASE FROM THE PLANT KINGDOM

FORESI, N. P.; CORREA ARAGUNDE, M.N.; CASALONGUE, C.; LAMATTINA, L.

San Miguel de Tucumán

Congreso; XLV Reunión Anual Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2009

SAIB

PL-P94. FIRST CHARACTERIZATION OF A NITRIC OXIDE SYNTHASE FROM THE PLANT KINGDOM Foresi NP; Correa-Aragunde MN; Casalongué CA; Lamattina L Instituto de Investigaciones Biológicas, FCEyN, UNMdP.CC 1245 (7600) Mar del Plata. E-mail: npforesi@mdp.edu.ar Recently, it has been completed the genome sequence of the unicellular green algae Ostreococcus tauri (Ot), the smallest member of the plant Kingdom. In the genome of Ot it has been identified a gene sharing high similarity with those coding for nitric oxide synthase (NOS) of mammals. NOS enzymes generate nitric oxide (NO) from arginine using NADPH as electron donor. NO is involved in many physiological processes and associated with responses to (a)biotic stresses in organisms from all Kingdoms. To date, no gene sequence, cDNA or protein coding for a putative plant NOS have been found. In this study, we have conducted the cloning, expression and biochemical characterization of OtNOS. The OtNOS has been successfully expressed as a recombiant protein in Escherichia coli. OtNOS was showed as high-spin heme protein and shared spectral properties to those described for others isolated NOS. Activity of the recombinant OtNOS was determined by two methods (i) hemoglobin capture and (ii) NADPH oxidation. Bacteria expressing the recombinant OtNOS display high levels of NO after supplementation with the NOS substrate L-arginine and were more resistant than wt to the oxidant hydrogen peroxide. All together, these results describe for the first time the presence of NOS in plant Kingdom. protein and shared spectral properties to those described for others isolated NOS. Activity of the recombinant OtNOS was determined by two methods (i) hemoglobin capture and (ii) NADPH oxidation. Bacteria expressing the recombinant OtNOS display high levels of NO after supplementation with the NOS substrate L-arginine and were more resistant than wt to the oxidant hydrogen peroxide. All together, these results describe for the first time the presence of NOS in plant Kingdom. member of the plant Kingdom. In the genome of Ot it has been identified a gene sharing high similarity with those coding for nitric oxide synthase (NOS) of mammals. NOS enzymes generate nitric oxide (NO) from arginine using NADPH as electron donor. NO is involved in many physiological processes and associated with responses to (a)biotic stresses in organisms from all Kingdoms. To date, no gene sequence, cDNA or protein coding for a putative plant NOS have been found. In this study, we have conducted the cloning, expression and biochemical characterization of OtNOS. The OtNOS has been successfully expressed as a recombiant protein in Escherichia coli. OtNOS was showed as high-spin heme protein and shared spectral properties to those described for others isolated NOS. Activity of the recombinant OtNOS was determined by two methods (i) hemoglobin capture and (ii) NADPH oxidation. Bacteria expressing the recombinant OtNOS display high levels of NO after supplementation with the NOS substrate L-arginine and were more resistant than wt to the oxidant hydrogen peroxide. All together, these results describe for the first time the presence of NOS in plant Kingdom. protein and shared spectral properties to those described for others isolated NOS. Activity of the recombinant OtNOS was determined by two methods (i) hemoglobin capture and (ii) NADPH oxidation. Bacteria expressing the recombinant OtNOS display high levels of NO after supplementation with the NOS substrate L-arginine and were more resistant than wt to the oxidant hydrogen peroxide. All together, these results describe for the first time the presence of NOS in plant Kingdom. Ostreococcus tauri (Ot), the smallest member of the plant Kingdom. In the genome of Ot it has been identified a gene sharing high similarity with those coding for nitric oxide synthase (NOS) of mammals. NOS enzymes generate nitric oxide (NO) from arginine using NADPH as electron donor. NO is involved in many physiological processes and associated with responses to (a)biotic stresses in organisms from all Kingdoms. To date, no gene sequence, cDNA or protein coding for a putative plant NOS have been found. In this study, we have conducted the cloning, expression and biochemical characterization of OtNOS. The OtNOS has been successfully expressed as a recombiant protein in Escherichia coli. OtNOS was showed as high-spin heme protein and shared spectral properties to those described for others isolated NOS. Activity of the recombinant OtNOS was determined by two methods (i) hemoglobin capture and (ii) NADPH oxidation. Bacteria expressing the recombinant OtNOS display high levels of NO after supplementation with the NOS substrate L-arginine and were more resistant than wt to the oxidant hydrogen peroxide. All together, these results describe for the first time the presence of NOS in plant Kingdom. protein and shared spectral properties to those described for others isolated NOS. Activity of the recombinant OtNOS was determined by two methods (i) hemoglobin capture and (ii) NADPH oxidation. Bacteria expressing the recombinant OtNOS display high levels of NO after supplementation with the NOS substrate L-arginine and were more resistant than wt to the oxidant hydrogen peroxide. All together, these results describe for the first time the presence of NOS in plant Kingdom. scherichia coli. OtNOS was showed as high-spin heme protein and shared spectral properties to those described for others isolated NOS. Activity of the recombinant OtNOS was determined by two methods (i) hemoglobin capture and (ii) NADPH oxidation. Bacteria expressing the recombinant OtNOS display high levels of NO after supplementation with the NOS substrate L-arginine and were more resistant than wt to the oxidant hydrogen peroxide. All together, these results describe for the first time the presence of NOS in plant Kingdom. Financed by Conicet, ANPCyT and UNMDP.