Urea Output by L3 Teladorsagia circumcincta and some Properties of Two Urea Producing Enzymes
-
N Muhamad
,
Lr Walker
,
Dc Simcock
,
Kc Pedley
,
Hv Simpson
,
S Brown
Abstract
Background: Like several other parasites, Teladorsagia circumcincta secretes or excretes urea, but nei-ther the rate of efflux nor the possible metabolic sources of the urea has been considered.
Methods: Parasites were maintained by passage through sheep. Urea efflux was measured using phe-nol/hypochlorite after treatment with urea aminohydrolase. The kinetics of creatine amidinohy-drolase and arginine amidinohydrolase were characterised by coupling the reactions with urea ami-nohydrolase and glutamate dehydrogenase.
Results: Infective L3 T. circumcincta secreted or excreted urea at 25% of the rate of NH3/NH4+. The rate of urea efflux was about 84 pmol h-1 (103 larvae)-1 over 4 hours, corresponding to about 11 nmol h-1 mg-1 protein. We could not detect urea aminohydrolase activity, but urea production by both creatine amidinohydrolase and arginine amidinohydrolase could be detected. The apparent Km and Vmax of creatine amidinohydrolase were 1.1 mM and 48 nmol h-1 mg-1 protein, respectively, and the activity was greatest at pH 8. The apparent Km and Vmax of arginine amidinohydrolase were 0.7 mM and 62 nmol h-1 mg-1 protein, respectively, and the activity was greatest at pH 7.9.
Conclusion: The activity of creatine amidinohydrolase and arginine amidinohydrolase was sufficient to account for the rate of urea secretion or excretion.
Wright DJ, Newall DR. Nitrogen excretion, osmotic and ionic regulation in nematodes. In: Croll NA, ed. The Organization of Nematodes. London: Academic Press Inc., London, Ltd, 1976:163-210.
Rogers WP. Nitrogen catabolism in nema-tode parasites. Aust J Biol Res. 1952; B5: 210-222.
Cavier RM, Savel J. Le métabolism proté-ique de l'Ascaris du porc, Ascaris lumbricoides Linné, 1758, est-il ammoniotélique ou uréotélique? C R Acad Sci. 1954; 238: 2448-2450.
Paltridge RW, Janssens PA. A reinvestiga-tion of the status of the ornithine-urea cycle in adult Ascaris lumbricoides. Comp Biochem Physiol. 1971; 40B: 503-513.
Muhamad N, Pedley KC, Simpson HV, Brown S. Ammonium excretion/secretion by L3 Ostertagia circumcincta and its connection with nitrogen metabolism. N Z J Zool. 2004; 31: 377.
Muhamad N, Walker LR, Pedley KC, Sim-cock DC, Brown S. The kinetics of NH3/NH4+ efflux from L3 Teladorsagia cir-cumcincta. Parasitol Int. 2012; 61: 487-492.
Simpson HV, Muhamad N, Walker LR, Simcock DC, Brown S, Pedley KC. Nitro-gen excretion by the sheep abomasal para-site Teladorsagia circumcincta. Exptl Parasitol. 2009; 123: 17-23.
Muhamad N, Brown S, Pedley KC, Simp-son HV. Some aspects of nitrogen metabo-lism in Ostertagia circumcincta. N Z J Zool. 2005; 32: 278-279.
Umair S, Patchett ML, Bland RJ, Simpson HV. Arginine metabolism in the sheep abomasal nematode parasites Haemonchus contortus and Teladorsagia circumcincta. Exptl Parasitol. 2011; 127: 506-514.
Fox BA, Gigley JP, Bzik DJ. Toxoplasma gondii lacks the enzymes required for de novo arginine biosynthesis and arginine starvation triggers cyst formation. Int J Parasitol. 2004; 34: 323-331.
Cook T, Roos D, Morada M, Zhu G, Keithly JS, Feagin JE, Wu G, Yarlett N. Di-vergent polyamine metabolism in the api-complexa. Microbiology. 2007; 153: 1123-1130.
Simcock DC, Brown S, Neale JD, Przemeck SMC, Simpson HV. L3 and adult Ostertagia (Teladorsagia) circumcincta exhibit cyanide sen-sitive oxygen uptake. Exptl Parasitol. 2006; 112: 1-7.
Walker LR, Simcock DC, Neale JD, Simp-son HV, Brown S. Thermotolerance of L3 Ostertagia (Teladorsagia) circumcincta and some of its enzymes. Vet Parasitol. 2007; 146: 77-82.
Bolleter WT, Bushman CJ, Tidwell PW. Spectrophotometric determination of am-monia as indophenol. Anal Chem. 1961; 33: 592-594.
Bradford MM. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976; 72: 248-254.
Sapan CV, Lundblad RL, Price NC. Col-orimetric protein assay techniques. Biotech-nol Appl Biochem. 1999; 29: 99-108.
Mora J, Tarrab R, Martuscelli J, Soberón G. Characteristics of arginases from ureotelic and non-ureotelic animals. Biochem J. 1965; 96: 588-594.
Coll M, Knof SH, Ohga Y, Messerschmidt A, Huber R, Moellering H, Rüssmann L, Schumacher G. Enzymatic mechanism of creatine amidinohydrolase as deduced from crystal structures. J Mol Biol. 1990; 214: 597-610.
Reczkowski RS, Ash DE. Rat liver arginase: kinetic mechanism, alternate substrates, and inhibitors. Arch Biochem Biophys. 1994; 312: 31-37.
Matsuda Y, Wakamatsu N, Inouye Y, Uede S, Hashimoto Y, Asano K, Nakamura S. Purification and characerization of creatine amidinohydrolase of Alcaligenes origin. Chem Pharm Bull. 1986; 34: 2155-2160.
Muhamad N, Simcock DC, Pedley KC, Simpson HV, Brown S. The kinetics of glu-tamate dehydrogenase of Teladorsagia circum-cincta and the lifestyle of the parasite. Comp Biochem Physiol. 2011; 159B: 71-77.
Ihaka R, Gentleman R. R: a language for data analysis and graphics. J Comput Graphical Stat. 1996; 5: 299-314.
Christianson DW. Arginase: structure, mechanism, and physiological role in male and female sexual arousal. Acc Chem Res. 2005; 38: 191-201.
Warner RC. The kinetics of the hydrolysis of urea and of arginine. J Biol Chem. 1942; 142: 705-723.
Yoshimoto T, Oka I, Tsuru D. Purification, crystallization, and some properties of creatine amidinohydrolase from Pseudomonas putida. J Biochem. 1976; 79: 1381-1383.
Yoshimoto T, Oka I, Tsuru D. Creatine amidinohydrolase of Pseudomonas putida: crys-tallization and some properties. Arch Bio-chem Biophys. 1976; 177: 508-515.
Schumann J, Böhm G, Schumacher G, Ru-dolph R, Jaenicke R. Stabilization of creati-nase from Pseudomonas putida by random mutagenesis. Protein Sci. 1993; 2: 1612-1620.
Kaplan A, Szabo LL. Creatinine hydrolase and creatine amidinohydrolase: II. Partial purification and properties. Mol Cell Bio-chem. 1974; 3: 17-25.
Weinland E. Über die Zersetzung stickstoff-haltiger Substanz bei Ascaris. Z Biol. 1904; 45: 517-531.
| Files | ||
| Issue | Vol 8 No 1 (2013) | |
| Section | Articles | |
| Keywords | ||
| Kinetics Teladorsagia circumcincta Urea | ||
| Rights and permissions | |
|
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License. |
