Glyceraldehyde-3-Phosphate Dehydrogenase in Greening Zea mays L. Leaves

Abstract

Nicotinamide adenine dinucleotide phosphate (NADP)-dependent glyceraldehyde-3-phosphate dehydrogenase (GPDH) (EC 1.2.1.13), a chloroplast enzyme, had low activity in etioplasts of maize leaves. A light dependent increase of enzyme activity of 7-day-old etiolated seedlings showed a lag period of about 2.5 hours followed by a rapid increase in activity during the next 10 hours. The chlorophyll content followed a similar pattern of increasing concentration, but its formation was not directly related to NADP-GPDH formation. The specific activity of NADP-GPDH was lowest in the morphologically youngest tissue near the base of the lamina. The increase in NADP-GPDH was inhibited by cycloheximide but not by chloramphenicol. This indicates that at least some of the enzyme polypeptides are synthesized by 80S ribosomes in the cytoplasm, transported into chloroplasts and become active in chloroplasts.In etiolated maize shoots subjected to a combination of both 3-(p-chlorophenyl)-1,1-dimethylurea, monuron at 7 x 10-i M and far red light treatment for 15 hours, the NADP-GPDH activity increased 42% over the dark control compared to 70% increase for the light control. It is concluded that NADPH is not absolutely required for the activation of NADP-GPDH in maize leaves under physiological conditions. In higher plants, NADPH-specific glyceraldehyde-3-P dehydrogenase (NADP-GPDH) (EC 1.2.1.13) occurs only in photosynthetic tissue (4,11,12,14). Seedlings grown in the dark contain little NADPH-GPDH activity, but the enzyme activity increases markedly if the seedlings are exposed to the light (14,19,20). A comprehensive investigation of the specificity of glyceraldehyde-3-P dehydrogenase was done by Pupillo (24) and Pupillo and Piccari (25) who concluded that in all green plants both the NADand NADP-dependent GPDH activities of chloroplasts are catalyzed by one enzyme which is different from the NAD-linked cytoplasmic enzyme. Schulman and Gibbs (26) with 2.7 mm NADPH gave up to a 600%o increase of NADP-GPDH activity over the control. This increase in the activity was accompanied by a decrease in the activity of the NAD-linked GPDH. The activation with NADPH was not a nonspecific reduction of disulfide linkages, since DTT could not substitute for NADPH in the activation by NADPH in the crude homogenate. NADPH was also reported to activate partially purified NADP-GPDH (23). The concentration curve response to NADPH activation was sigmoidal in shape. It was postulated that in the light there is an increased concentration of NADPH and that the NADPH allosterically activates the GPDH causing a transition from a NAD-dependent to a NADP-dependent form of the enzyme (23). Pupillo and Piccari (25) were able to obtain an equilibrium mixture of different oligomers in several discrete aggregational states depending upon the experimental conditions. The protomer, which has a low Km for NADP+ and NAD+, reverts to tetramers in the absence of NADP+ with considerable increase in the apparent Km for NADP+. NAD+ was unable to induce dissociati...

Proteomic Analysis of Somatic Embryogenesis in Cyclamen persicum Mill

“…The detection of triosephosphate isomerase in somatic embryogenesis of Cyclamen in this study is similar to those findings reported in various species (Milena et al 2008;Nogueira et al 2007;Winkelmann et al 2006). Moreover, protein spot 24, a nicotinamide adenine dinucleotide phosphate (NADP)-dependent glyceraldehyde-3-phosphate dehydrogenase, is a chloroplast enzyme (Chin and Ralph 1980) and is activated by light (Ziegleir et al 1965). This protein may be involved in photosynthetic electron transport and it is correlated to active proliferation of global somatic embryogenesis requiring energy.…”

A starch‐accumulating mutant ofArabidopsis thalianadeficient in a chloroplastic starch‐hydrolysing enzyme

“…Glyceraldehyde‐3‐phosphate dehydrogenase: The 1 ml reaction mixture contained 50 m m glycylglycine, pH 8.2, 80 m m 3‐phosphoglyceric acid (3‐PGA), 20 m m MgSO 4 , 10 m m cysteine, 5 m m glutathione (reduced form), 2.5 m m ATP, 0.4 m m NADPH and 2 units 3‐phosphoglycerate kinase (from yeast) ( Lin & Stocking 1980).…”

Characterisation of pea cytosolic glutathione reductase expressed in transgenic tobacco

“…Each assay datum point is the mean of three determinations. The chloroplast marker enzyme was glyceraldehyde-3-phosphate dehydrogenase (GAPDH; Lin and Stocking 1980), the cytosol marker enzyme was UDP-glucose pyrophosphorylase (UDPGPP; Smith et al 1989). …”

“…Marker enzyme used for the chloroplast was NADP-linked glyceraldehyde-3-phosphate dehydrogenase (GAPDH) assayed by the oxidation of NADPH (Lin and Stocking 1980). The cytosolic marker enzyme used was UDP-glucose pyrophosphorylase (Smith et al 1989).…”