Urea Is a Product of Ureidoglycolate Degradation in Chickpea. Purification and Characterization of the Ureidoglycolate Urea-Lyase

Abstract

A ureidoglycolate-degrading activity was analyzed in different organs of chickpea (Cicer arietinum). Activity was detected in all the tissues analyzed, but highest levels of specific activity were found in pods, from which it has been purified and characterized. This is the first ureidoglycolate-degrading activity that has been purified to homogeneity from any photosynthetic organism. Only one ureidoglycolate-degrading activity was found during the purification. The enzyme was purified 1,500-fold, and specific activity for the pure enzyme was 8.6 units mg Ϫ1 , which corresponds with a turnover number of 1,600 min Ϫ1 . The native enzyme has a molecular mass of 180 kD and consists of six identical or similar-sized subunits of 31 kD each. The enzyme exhibited hyperbolic, Michaelian kinetics for (Ϫ) ureidoglycolate with K m values of 6 and 10 m in the presence or absence of Mn 2ϩ , respectively. Optimum pH was between 7 and 8 and maximum activity was found at temperatures above 70°C, the enzyme being extremely stable and resistant to heat denaturation. The activity was inhibited by EDTA and enhanced by several bivalent cations, thus suggesting that the enzyme is a metalloprotein. This enzyme has been characterized as a ureidoglycolate urea-lyase (EC 4.3.2.3), which catalyzes the degradation of (Ϫ) ureidoglycolate to glyoxylate and urea. This is the first time that such an activity is detected in plant tissues. A possible function for this activity and its implications in the context of nitrogen mobilization in legume plants is also discussed.

The Ureide-Degrading Reactions of Purine Ring Catabolism Employ Three Amidohydrolases and One Aminohydrolase in Arabidopsis, Soybean, and Rice

“…Especially controversial is the nature of the ureidoglycolate-degrading reaction. From chickpea (Cicer arietinum) and common bean (Phaseolus vulgaris), enzymes were purified that, in contrast to UAH, cleave ureidoglycolate to glyoxylate and urea (Muñoz et al, 2001(Muñoz et al, , 2006. Consistently, it was reported that in soybean, half of the nitrogen from allantoate was released as urea, presumably during ureidoglycolate degradation (Todd and Polacco, 2004).…”

Nickel Deficiency Disrupts Metabolism of Ureides, Amino Acids, and Organic Acids of Young Pecan Foliage

“…UGL or UGAH), with one predominating over the other. For example, in soybean, ureidoglycolate catabolism was approximately 95% via UGAH and approximately 5% via UGL; thus, urease did not appear to be essential for ureide catabolism in this species (Stebbins and Polacco, 1995), whereas in chickpea (Cicer arietinum) UGL appears to be the predominant path (Munoz et al, 2001). The accumulation of ureidoglycolate and quantitative reduction in urea from Ni-D foliage suggests that the UGL activity is being impaired.…”

“…Additionally, the accumulation of ureidoglycolate also suggests either that pecan might not possess the second pathway for the catabolism of ureidoglycolate or that the activity of UGAH is also being impaired by Ni deficiency. Ureidoglycolase is a metalloenzyme whose activity is inhibited by EDTA and enhanced by several different bivalent cations (Munoz et al, 2001). It is possible that one of these metal components is Ni.…”

“…The accumulated ureidoglycolate is potentially degraded by two enzymes (Wells and Lees, 1991;Munoz et al, 2001). One is via ureidoglycolate amidinohydrolase (UGL; EC 4.3.2.3) to produce urea and glyoxylate.…”

“…One is via ureidoglycolate amidinohydrolase (UGL; EC 4.3.2.3) to produce urea and glyoxylate. The second is via ureidoglycolate amidohydrolase (UGAH; EC 3.5.3.19) to produce NH 4 , CO 2 , and glyoxylate (Wells and Lees, 1991;Munoz et al, 2001), and requires manganese as a cofactor (Lukaszeski et al, 1992;Vadez and Sinclair, 2000).…”