Glutathione and Homoglutathione Synthesis in Legume Root Nodules

Abstract

High-performance liquid chromatography (HPLC) with fluorescence detection was used to study thiol metabolism in legume nodules. Glutathione (GSH) was the major non-protein thiol in all indeterminate nodules examined, as well as in the determinate nodules of cowpea (Vigna unguiculata), whereas homoglutathione (hGSH) predominated in soybean (Glycine max), bean (Phaseolus vulgaris), and mungbean (Vigna radiata) nodules. All nodules had greater thiol concentrations than the leaves and roots of the same plants because of active thiol synthesis in nodule tissue. The correlation between thiol tripeptides and the activities of glutathione synthetase (GSHS) and homoglutathione synthetase (hGSHS) in the nodules of eight legumes, and the contrasting thiol contents and activities in alfalfa (Medicago sativa) leaves (98% hGSH, 100% hGSHS) and nodules (72% GSH, 80% GSHS) indicated that the distribution of GSH and hGSH is determined by specific synthetases. Thiol contents and synthesis decreased with both natural and induced nodule senescence, and were also reduced in the senescent zone of indeterminate nodules. Thiols and GSHS were especially abundant in the meristematic and infected zones of pea (Pisum sativum) nodules. Thiols and ␥-glutamylcysteinyl synthetase were also more abundant in the infected zone of bean nodules, but hGSHS was predominant in the cortex. Isolation of full-length cDNA sequences coding for ␥-glutamylcysteinyl synthetase from legume nodules revealed that they are highly homologous to those from other higher plants.The tripeptide glutathione (GSH; ␥Glu-Cys-Gly) is the major non-protein thiol in most animals, plants, and prokaryotes (Meister and Anderson, 1983; Hausladen and Alscher, 1993; Rennenberg, 1997). In plants, GSH is a versatile antioxidant that can directly scavenge activated oxygen species and participate in the ascorbate-GSH cycle for peroxide removal in the chloroplasts. It is also involved in many other vital functions of plants, including the transport and storage of sulfur, the synthesis of proteins and DNA, tolerance to abiotic and biotic stress, and the detoxification of xenobiotics, air pollutants, and heavy metals (Hausladen and

Posttranslational control of transcription factor FixK 2 , a key regulator for the Bradyrhizobium japonicum –soybean symbiosis

“…To demonstrate such effects experimentally will be difficult, even more so as we did not notice a phenotypic difference between the WT and the C183A-FixK 2 mutant in terms of nodule formation and nitrogen fixation activity. (ii) Although mature nodules are thought to be relatively well protected from ROS by enzymes from both symbiotic partners (37), senescent nodules enhance ROS production and concurrently decrease the antioxidant defense (38,39), leading to oxidative damage of lipids, proteins, and DNA (40). Again, ROS-dependent shut-down of FixK 2 activity would assist in down-regulating symbiotic functions that become futile during senescence.…”

Structural Basis for Evolution of Product Diversity in Soybean Glutathione Biosynthesis

“…Nearly all eukaryotes and prokaryotes synthesize the multifunctional peptide GSH (Meister, 1995); however, some plants also synthesize GSH analogs with substitutions of the terminal Gly ( Figure 1A) (Rauser et al, 1986;Klapheck et al, 1994;Klapheck et al, 1995;Meuwly et al, 1995;Matamoros et al, 1999). In particular, many legumes produce hGSH for root nodulation (Matamoros et al, 2003;Frendo et al, 2005;Loscos et al, 2008).…”

“…In nearly all organisms, glutathione synthetase (GS) catalyzes the addition of Gly to gEC (Meister, 1995;Herrera et al, 2007). In legumes, homoglutathione synthetase (hGS) uses b-Ala instead of Gly to form hGSH (Matamoros et al, 1999;Frendo et al, 2001;Iturbe-Ormaetxe et al, 2002). Although GS and hGS share similar reaction mechanisms based on biochemical and structural studies, the molecular basis for the difference in substrate specificity is unclear due to no available structural data for any plant GS or hGS.…”

“…Although GSH is the predominant thiol-containing tripeptide found in plants, various plant species produce glutathione homologs in which the terminal Gly is substituted with a different amino acid ( Figure 1A). For example, legumes make GSH, in addition to producing homoglutathione (hGSH), in which b-Ala replaces Gly, in a tissue-specific manner (Klapheck et al, 1995;Matamoros et al, 1999). Synthesis of hGSH maintains redox balance in legume nodules (Moran et al, 2000) and is critical for rhizobia-legume nodulation in roots (Matamoros et al, 2003;Frendo et al, 2005;Loscos et al, 2008).…”

Nodules and oxygen

“…Furthermore, O 2 can be reduced directly by nitrogenase, hydrogenase and ferredoxin in the bacteroids (Dalton 1995). Not surprisingly, legume nodules contain high activities of the ascorbateglutathione cycle enzymes (Matamoros et al 1999a; and millimolar concentrations of ascorbate (Matamoros et al 1999b).…”