Arginine and Ornithine Decarboxylases, the Polyamine Biosynthetic Enzymes of Mung Bean Seedlings

Abstract

General properties and relative activities of L-arginine decarboxylase (ADC) (EC 4.1.1.19) and L-ornithine decarboxylase (ODC) (EC 4.1.1.17), two important enzymes in putrescine and polyamine biosynthesis, were investigated in mung bean (Vigna radiata L.) tissues. Both activities increase linearly with increasing concentrations of crude enzyme, but the increase in ADC activity is considerably greater. The decarboxylation reaction is linear for up to 30 to 60 minutes, and both enzymes have a pH optimum of 7.2. a-Difluoromethyl-ornithine inhibits ODC activity of excised roots, while increasing ADC activity.High specific activity of both enzymes is detected in terminal buds and leaves, while root and hypocotyl activity is low. Different ADC-to-ODC activity ratios are found in various tissues of mung bean plants. Substantial increase in the activity of both enzymes is detected in incubated sections as compared with intact plants. A The importance of polyamines in various growth and physiological processes in plants has been inferred from both application of exogenous polyamines and from changes in endogenous polyamines and related metabolites (1,2,4,9,13 2Abbreviations: ADC, L-arginie decarboxylase; ODC, L-ormithine de-carboxylase.ADC and ODC are active and involved in polyamine biosynthesis (16,19,21). In recent studies (10), ODC activity of developing tomato fruits was found to increase dramatically up to the 3rd day after pollination, while ADC activity was very low, and both arginine and ornithine were equally well decarboxylated in tobacco cell cultures (5).In view of the contradictory results regarding the activity and importance of ADC and ODC in plants, a comparative study of these two polyamine-biosynthetic enzymes was undertaken. We report the kinetics and several properties of ADC and ODC as well as their relative activity in tissues of mung bean seedlings.MATERIALS AND METHODS Plant Material. Mung bean (Vigna radiata L.) seeds were germinated and grown in vermiculite in a controlled phytotron (27°C-day and 22°C-night temperature, 16-h photoperiod, 78% RH). One cm subapical sections of epicotyls were harvested from 7-to 8-day-old seedlings and were extracted for determination of enzyme activities. Occasionally, other plant parts were extracted, as mentioned. Epicotyl sections of French bean (Phaseolus vulgaris L. var. Brittle Wax) and cotton (Gossypium hirsutum L. SJ-2), grown for 8 days under the same conditions, were also used in several experiments.Extraction and Assay of ADC and ODC. Sections were harvested, weighed, and ground in a prechilled mortar with a pestle (250-600 mg fresh weight/2 ml medium). Extraction medium consisted of 10 mm phosphate buffer (pH 7.2), 0.1 mm DTT, 1 mm pyridoxal-5'-phosphate, and 20 mm Na-EDTA. The extracts were centrifuged at 12,000g for 20 min, and the supernatant, hereafter referred to as crude enzyme, was used immediately.

Assaying Ornithine and Arginine Decarboxylases in Some Plant Species

“…The choice of pH 8.0 buffers for the plant enzyme assays was based on the optima reported for ODC and ADC, which range from 7.2 to 8.5 with Tris-HCI buffer being used most frequently (1,7,15,16,18). The activities of ODC and ADC in various plant species show an extremely broad range; they were determined at extremely different substrate concentrations ranging from 14 uM to 5 mM of L-Orn or L-Arg (1,14,16,27) 'The aliquots used corresponded to 40 mg fresh per reaction mixture.…”

“…The activities of ODC and ADC in various plant species show an extremely broad range; they were determined at extremely different substrate concentrations ranging from 14 uM to 5 mM of L-Orn or L-Arg (1,14,16,27) 'The aliquots used corresponded to 40 mg fresh per reaction mixture. The distribution of radioactivity in the blanks did not differ depending on the buffer.…”

The effects of spermidine synthesis inhibitors on in-vitro plant development

“…Ornithine decarboxylase (ODC), which catalyses the formation of putrescine from omithine, has an important role in the growth of selected plant species [1,3,9] and may be essential for embryo development in animals [6]. Elevated levels of polyamines and arginine decarboxylase (ADC), the enzyme involved in the alternate pathway of putrescine biosynthesis from arginine in plants, have been associated with embryogenesis in cultures of wild carrot [ 15,16].…”

Variations in ergosterol content and ornithine decarboxylase activity of ectomycorrhizal root-soil systems

“…As the ODC assay includes root and fungal sources, work should continue in pure roots, perhaps monitoring ODC in conjunction with other biochemical techniques such as arginine decarboxylase (ADC) found in significant levels in roots (Altman et al, 1982). This enzyme may be used in root bioactivity determinations as most microoganisms do not favor the ADC pathway for polyamine production (Cohen,197l).…”