Theses Doctoral

PhD student:

Ana Salas Huertas


María Jesús Delgado Igeño

Date of presentation:


Faculty and University:

Facultad de Ciencias, Universidad de Granada

Qualification grade:

Sobresaliente cum laude


Exploring the role of the haemoglobin from Bradyrhizobium diazoefficiens in nitric oxide detoxification during free-living and endosymbiotic lifestyles


In this PhD Thesis, the in vitro role of Bjgb haemoglobin and the flavoprotein Flp from B. diazoefficiens in NO metabolism has been demonstrated. The ability of Flp to reduce Bjgb has been demonstrated. Once Bjgb is reduced, the heme group of this protein is able to bind NO. With these results, the ability of Bjgb to bind NO in vitro has been demonstrated, which would confirm the role of this protein to detoxify NO in vivo.
In this PhD thesis, we have also studied the involvement of Bjgb from B. diazoefficiens in the symbiotic interaction with soybean plants and in NO homeostasis in nodules. Using the 15N isotope dilution technique and analysis of nitrogenase enzyme activity and expression, it has been shown that inoculation of soybean plants with the Bjgb mutant strain confers tolerance to waterlogging. This effect is due to the reduction of NO accumulation in the nodules which is due to the induction of the expression and activity of nitric oxide reductase (Nor), which is the main protein involved in the elimination of NO in soybean nodules.
Finally, the involvement in symbiosis of the NO3- assimilation process has been studied. By analysing the nitrogenase activity and leghemoglobin (Lb) content of soybean nodules, it has been shown that assimilative nitrate reductase (NasC) does not play a relevant role in the reduction of NO3- in bacteroids, being periplasmic nitrate reductase (NapA) the main enzyme involved.
The results obtained during this PhD Thesis have contributed to increase the knowledge about the in vivo and in vitro function of the Bjgb protein of B. diazoefficiens in NO detoxification, both in free life and in symbiotic association with soybean plants. In addition, the possible role of the NasC and NirA proteins of B. diazoefficiens in symbiosis with soybean plants has been established.