Identification and characterization of proteins involved in rice urea and arginine catabolism.
Rice (Oryza sativa) production relies strongly on nitrogen (N) fertilization with urea, but the proteins involved in rice urea metabolism have not yet been characterized. Coding sequences for rice arginase, urease, and the urease accessory proteins D (UreD), F (UreF), and G (UreG) involved in urease activation were identified and cloned. The functionality of urease and the urease accessory proteins was demonstrated by complementing corresponding Arabidopsis (Arabidopsis thaliana) mutants and by multiple transient coexpression of the rice proteins in Nicotiana benthamiana. Secondary structure models of rice (plant) UreD and UreF proteins revealed a possible functional conservation to bacterial orthologs, especially for UreF. Using amino-terminally StrepII-tagged urease accessory proteins, an interaction between rice UreD and urease could be shown. Prokaryotic and eukaryotic urease activation complexes seem conserved despite limited protein sequence conservation for UreF and UreD. In plant metabolism, urea is generated by the arginase reaction. Rice arginase was transiently expressed as a carboxyl-terminally StrepII-tagged fusion protein in N. benthamiana, purified, and biochemically characterized (K(m) = 67 mm, k(cat) = 490 s(-1)). The activity depended on the presence of manganese (K(d) = 1.3 microm). In physiological experiments, urease and arginase activities were not influenced by the external N source, but sole urea nutrition imbalanced the plant amino acid profile, leading to the accumulation of asparagine and glutamine in the roots. Our data indicate that reduced plant performance with urea as N source is not a direct result of insufficient urea metabolism but may in part be caused by an imbalance of N distribution.
Cao
Feng-Qiu
FQ
Key Laboratory of Plant and Soil Interactions, College of Resources and Environmental Sciences, China Agricultural University, Beijing, China.
Werner
Andrea K
AK
Dahncke
Kathleen
K
Romeis
Tina
T
Liu
Lai-Hua
LH
Witte
Claus-Peter
CP
D020121:5' Untranslated Regions / Q000235:genetics; D001051:Apoenzymes / Q000378:metabolism; D017360:Arabidopsis / Q000187:drug effects / Q000201:enzymology / Q000235:genetics; D001119:Arginase / Q000378:metabolism; D001120:Arginine / Q000378:metabolism*; D003001:Cloning, Molecular; D018506:Gene Expression Regulation, Plant / Q000187:drug effects; D005816:Genetic Complementation Test; D018525:Germination / Q000187:drug effects; D007438:Introns / Q000235:genetics; D008969:Molecular Sequence Data; D009566:Nitrates / Q000494:pharmacology; D012275:Oryza / Q000187:drug effects / Q000201:enzymology / Q000254:growth & development / Q000378:metabolism*; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding / Q000187:drug effects; D000644:Quaternary Ammonium Compounds / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D014508:Urea / Q000378:metabolism* / Q000494:pharmacology; D014510:Urease / Q000737:chemistry / Q000235:genetics
D020121:5' Untranslated Regions; D001051:Apoenzymes; D009566:Nitrates; D010940:Plant Proteins; D000644:Quaternary Ammonium Compounds; D012333:RNA, Messenger; D014508:Urea; D001120:Arginine; D014510:Urease; D001119:Arginase