Nucleotide Sequence of a cDNA Encoding a Soybean Seedling Axes Arginase (Accesion No. AF035671).

ARIEL GOLDRAIJ; PATRICIA COELLO; JOE POLACCO

PLANT PHYSIOLOGY.

American Society of Plant Biologists

Lugar: Rockville, MD,USA; Año: 1998 vol. 116 p. 867 - 867

0032-0889

Arginine is an important nitrogen transport and storage compound which plays a critical role in the nitrogen economy of many plants. In soybean, arginine accounts for 18 % of the total seed protein nitrogen and up to 62 % of free amino acid nitrogen in developing embryos is in the arginine pool (Micallef and Shelp, 1989a). The first step of arginine degradation is catalyzed by arginase (L-Arg amidinohydrolase, EC 3.5.3.1), rendering urea and L-Ornithine. The nitrogen contained in urea is recycled to ammonia by urease action (Polacco and Holland, 1993) and ornithine is a precursor of proline, glutamic acid and polyamines. While arginine is actively synthesized in developing soybean cotyledons, arginase has been postulated to be inoperative in vivo (Stebbins et al, 1991) or to hydrolyze a minor portion of the large free arginine pool (Micallef and Shelp, 1989b). In contrast developing soybean cotyledon arginase is easily measured in vitro. Arginase activity increases sharply during germination (Kang and Cho, 1990) in which the products of its action can act as nitrogen and carbon skeleton sources to the growing axis. Considerable accumulation of urea derived from arginine has been observed in seedlings of a urease-negative soybean mutant (Stebbins et al, 1991; Stebbins and Polacco, 1995). Our primary interest is to study the regulation of arginase activity in developing and germinating embryos, determining first if arginases of these stages are distinct enzymes since this may explain differences observed in in vivo activities. A full length cDNA clone encoding a seedling arginase was recovered from an axes cDNA library in the lambda ZAP II vector by screening with an Arabidopsis arginase cDNA (Krumpelman et al, 1995; accesion number U15019). The cDNA sequence contains 1324 bp and at least two open reading frames that encode 350 and 327 aminoacids whose predicted masses are 38.6 and 35.9 kDa respectively, different from the size (60 kDa) previously reported (Kang and Cho, 1990) but in agreement with the average size of the arginase subunit from the major phylogenetic groups (Jenkinson et al, 1996). One of the three domains with a histidine conserved in several species, and proposed to be a Mn (II) ligand (Cavalli et al, 1994), is present in soybean arginase. A search with DNASIS sequence analysis software (Hitachi Software Engineering Co) showed that soybean arginase is 78 % identical to Arabidopsis arginase at the aminoacid level. Interestingly, the major divergence is in the 40 aminoacids of the N-terminal region. The most abundant amino acids of this region are Arg, Lys, Ser, Ala (each at 10%) and Leu (8%). A secondary structure prediction program indicates four domains with the ability to form alpha helical structures. Both helicity and an abundance of the indicated aminoacids are common in N-terminal signal peptides (Claros and Vicens, 1996). In agreement, all plant arginases studied so far have been reported to be mitochondrial or particulate enzymes (Polacco and Holland, 1993).