Kinetic Properties of Phosphoenolpyruvate Carboxylase in Peperomia camptotricha

Abstract

Kinetic characteristics of phosphoenolpyruvate carboxylase (PEPC) from the epiphytic C3 or C4: CAM intermediate plant, Peperomia camptotricha, were investigated. Few day versus night differences in V,X,K,(PEP), or malate inhibition were observed, even in extracts from water-stressed plants which characteristically perform CAM, regardless of efforts to stabilize day/night forms. The PEPC extracted from plants during the light period remained stable, without much of an increase or decrease in activity for at least 22 hours at 0 to 40C. Extracts from mature, fully developed leaves had slightly greater PEPC activity than from very young, developing leaves. Generally, however, the kinetic properties of'PEPC extracted from mature leaves of plants grown under short day (SD), long day (LD), or 1-week waterstress conditions, as well as from young, developing leaves, were similar. The PEPC inhibitor, L-malate, decreased the V.., and increased the K,,xpEp) for all treatments. Under specific conditions, malate did not inhibit PEPC rates in the dark extracts as much as the light. The PEPC activator, glucose-6-phosphate (G-6-P), lowered the K,,,p) for all treatments. At saturafing PEP concentrations, PEPC activity was independent of pH in the range of 7.5 to 9.0. At subsaturating PEP concentrations, the pH optimum was 7.8. The rates of PEPC activity were lower in the light period extracts than the dark, at pH 7.1, but day/night PEPC was equally active at pH 7.8. At pH 7.5 and a subsaturating PEP concentration, G-6-P significantly activated PEPC. At pH 8, however, only slight activation by G-6-P was observed. The lower pH of 7.5 combined with L-malate addition, greatly inhibited PEPC, particularly in extracts from young, developing leaves which were completely inhibited at an L-malate concentration of I millimolar. However, malate did not further inhibit PEPC activity in mature leaves when assayed at pH 7.1. The fairly constant day/night kinetic and regulatory properties of PEPC from P. camptotricha are unlike those of PEPC from CAM or C4 species studied, and are consistent with the photosynthetic metabolism of this plant.PEPC' is the initial C02-fixing enzyme in C4 plants (in the light) and CAM plants (in the dark). In both C4 and CAM plants, the amount of PEPC protein does not change between '

Kinetic characterization of phosphoenolpyruvate carboxylase extracted from whole-leaf and from guard-cell protoplasts of Vicia faba L. (C3 plant) with respect to tissue pre-illumination

“…(This enzyme fixes CO2 that is subsequently reduced (Melzer & O'Leary, 1987) and in this way functions like the so-called photosynthetic enzymes.) Until recently (Van Quy et at., 1991;Van Quy & Champigny, 1992), no evidence of light activation of this PEPC was reported (Chastain & Chollet, I989;Sipes & Ting, 1989).…”

Effect of Severe Water Stress on Aspects of Crassulacean Acid Metabolism in Xerosicyos

“…It is possible that the higher V,,, and lower K,(PEP) are in response to the decrease in tissue malate concentrations that result iinder water stress, since it is well known that in in vitro assays malate decreases the V,,, and increases the K,(PEP) (Kluge and Ting, 1978). However, we do not think that these observations are affected by an assay artifact, since desalting and dilution of the assay preparations in our experiments would reduce malate to levels far below the millimolar concentrations necessary to alter kinetic parameters (Sipes and Ting, 1989). Other possible effectors (Bandarian et al, 1992) would also be in low concentration.…”

“…PEPc activity was determined spectrophotometrically as previously described (Sipes and Ting, 1989). Leaf samples for assay were collected at the same time in the aftemoon.…”

Ontogenetic Development of Crassulacean Acid Metabolism as Modified by Water Stress in Peperomia

“…hatched bars water-stressed plants. Assays were conducted as previously described (Sipes and Ting 1989) and data were statistically obtained from rate curves (Canellas and Wedding 1980). Values are means±SD of three samples PEPC protein in P. scandens also increased with increasing age of leaves ( Fig.…”