Pyruvate Orthophosphate Dikinase of C₃ Seeds and Leaves as Compared to the Enzyme from Maize

Abstract

Pyruvate orthophosphate dikinase (PPDK) was found in varis immature seeds of C3 planRts (wheat, pea, green bean, plm, ad castor bean) in some C3 eaves (tobacco, spinach, sunflower, an wheat and in C (maize) kernels. The Protein Blot. PPDK was isolated from maize and purified as previously described (29). Antiserum to PPDK was prepared by injecting New eland white rabbits with 150 pg of the purified enzyme and Freund's complete adjuvant, followed after 26 d by a booster containing the same amount of enzyme and Freund's incomplete adjuvant and a second such booster at 40 d. The rabbits were bled at 41 d. The crude antiserum was used for the protein blot. Nonimmunized serum showed no cross-reaction with PPDK, and PPDK antiserum cross-reacted only with PPDK (1). This antiserum was used as a probe for the presence ofPPDK in.the various tissues studied.Each plant tissue studied (3 g) was ground in 6 ml of 0.1 M Tris buffer, pH 7.4, containing 10 mM MgC2, 18% w/v sucrose, and 1% P-mercaptoethanol. The homogenate was filtered and then centrifuged at 15,000g and OC for 20 min. Total soluble protein was determined by the method of Bradford (5). Crossreacting protein subunits similar in size to that of maize PPDK were detected by the protein blot method (6, 21).SDS-PAGE of each plant extract was carried out in a gradient gel (6.4 to 12.8%). Protein was transferred to cyanogen bromide paper prepared by the method of Clarke et al. (6). The tansfer paper was probed with anti-PPDK serum and then with J125 protein A. An autoradiograph was prepared using Kodak AR5 x-ray film with an intensifying screen at -70°C overnight.Densitometry. Relative levels of PPDK in preparations from different plant samples were estimated by densitometry of the xray film by comparison of the peak areas.

Low‐oxygen stress and water deficit induce cytosolic pyruvate orthophosphate dikinase (PPDK) expression in roots of rice, a C3 plant

“…A PEPC mediated CO 2 -fixation is known to be part of the metabolic response of plants during phosphorous deficiency, which also affects the energy-metabolism (Theodorou and Plaxton, 1993). Cytosolic PPDK has also been proposed to be involved in amino acid interconversions in seeds (Aoyagi and Bassham, 1984) and the recycling of coenzymes, which could be linked to the accumulation of proline in rice roots during low-oxygen stress (Bertani and Reggiani, 1991).…”

Inheritance of C4 Enzymes Associated with Carbon Fixation in Flaveria Species

“…The first relates to differences in stainability between polypeptides which vary significantly in structure. Although there is biochemical evidence suggesting that isoforms of the enzymes examined here are different in C3, C4, and C3-C4 plants, there is also evidence indicating that considerable structural integrity has been maintained throughout their divergence (2,14). As a result, differences stained nitrocellulose filter after transfer of gel as in A loaded with 75 sg protein/lane.…”

Pyruvate Orthophosphate Dikinase mRNA Organ Specificity in Wheat and Maize

“…In C4 leaves, PPDK2 is found in the chloroplasts of the mesophyll cells (8). PPDK is found also in seeds (1,12) and leaves (1) of wheat and in some other C3 plant tissues (for references, see 2). The role of PPDK in C3 leaf tissues is not clearly known, although it has been suggested that it might be involved in the control of stomatal opening (16).…”

“…The role of PPDK in C3 leaf tissues is not clearly known, although it has been suggested that it might be involved in the control of stomatal opening (16). The occurrence of PPDK in developing wheat seeds does not correlate quantitatively with photosynthetic enzymes (2), and it was suggested that one role of the enzyme might be in the conversion of C3 amino acids, especially alanine, to C4 and C5 amino acids.…”

“…similar mechanism may be present in leaves of wheat, a C3 plant. Previously, using protein blot analysis with the same maize antibody, we detected PPDK polypeptides no larger than 94 kD among proteins extracted from maize leaves and kernels and from wheat leaves and seeds (2).…”