Ribulose-1,5-Bisphosphate Carboxylase/Oxygenase Content,
Assimilatory Charge, and Mesophyll Conductance in Leaves |
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| Authors: | Hillar Eichelmann and Agu Laisk |
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| Affiliation: | Tartu Ülikooli Molekulaar-ja Rakubioloogia Instituut, Riia tn 23, Tartu, 51010, Estonia |
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| Abstract: | The content of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) (Et; EC 4.1.1.39) measured in different-aged leaves of sunflower (Helianthus annuus) and other plants grown under different light intensities, varied from 2 to 75 μmol active sites m−2. Mesophyll conductance (μ) was measured under 1.5% O2, as well as postillumination CO2 uptake (assimilatory charge, a gas-exchange measure of the ribulose-1,5-bisphosphate pool). The dependence of μ on Et saturated at Et = 30 μmol active sites m−2 and μ = 11 mm s−1 in high-light-grown leaves. In low-light-grown leaves the dependence tended toward saturation at similar Et but reached a μ of only 6 to 8 mm s−1. μ was proportional to the assimilatory charge, with the proportionality constant (specific carboxylation efficiency) between 0.04 and 0.075 μm−1 s−1. Our data show that the saturation of the relationship between Et and μ is caused by three limiting components: (a) the physical diffusion resistance (a minor limitation), (b) less than full activation of Rubisco (related to Rubisco activase and the slower diffusibility of Rubisco at high protein concentrations in the stroma), and (c) chloroplast metabolites, especially 3-phosphoglyceric acid and free inorganic phosphate, which control the reaction kinetics of ribulose-1,5-bisphosphate carboxylation by competitive binding to active sites.Rubisco (EC 4.1.1.39) catalyzes the irreversible carboxylation of RuBP to form two PGA molecules (in this work the oxygenase reaction was not active since a low O2 concentration was used). RuBP carboxylation is the major rate-determining reaction in photosynthetic CO2 assimilation. All factors that influence the photosynthetic rate do so by influencing the activity of Rubisco and the concentration of its substrates, CO2 and RuBP. Et in leaves may be as high as 75 μmol m−2, and for the extracted enzyme Km(CO2) = 9.4 μm (Makino et al., 1985a) and Km(RuBP) = 30 to 40 μm (Yeoh et al., 1981). In leaves photosynthesizing under atmospheric conditions, the concentration of RuBP may increase to 10 to 15 mm (Badger et al., 1984; Sharkey et al., 1986), but the concentration of CO2 is usually about 4 to 8 μm in leaf intercellular spaces, depending on stomatal conductance. This CO2 concentration is well below the Km(CO2) of the enzyme, and it is the initial slope of the kinetic curve VM/Km(CO2), termed carboxylation conductance, that becomes important.rc limits the CO2-fixation rate in series with the other resistances, rg and rmd. The carboxylation rates are usually expressed in relation to Ci or Cw. Cc is usually about 20% to 30% lower than Cw because of concentration decrease generated by the carboxylation flux on rmd. Considering the above, the carboxylation conductance in intact leaves in vivo may be found as the initial slope of the A versus Cc graph at low Cc values. If Cc cannot be calculated because rmd is unknown, the closest approximation is a plot of A versus Cw or A versus Ci. The true parameters of the carboxylase can be found only from experiments carried out in nonphotorespiratory conditions (1%–2% O2); otherwise the competing oxygenase reaction consumes a part of RuBP and partially inhibits carboxylase activity.Because of technical problems with the measurement of A versus Cw relationships, in many studies only the net photosynthetic rate under atmospheric conditions (21% O2) was related to Rubisco activity or content. Nevertheless, good correlation has been found (Makino et al., 1983; Hudson et al., 1992; Jacob and Lawlor, 1992; Jiang and Rodermel, 1995; Nakano et al., 1997). These results indicated that the level of Rubisco protein could be a limiting factor in photosynthesis throughout the life span of the leaf under natural environmental conditions. On the other hand, when Rubisco levels in leaves exceeded 4 g m−2 (60 μmol m−2), the in vivo Rubisco activity (measured as photosynthesis under pCi = 20 to 30 Pa and 21% O2) became curvilinearly correlated with Et (Makino et al., 1994, 1997). When measurements were made over the whole life span of wheat leaves, the measured rates of photosynthesis were lower in young leaves, which had high protein content, than would have been expected from the amount and activity of Rubisco (Lawlor et al., 1989).During senescence the decrease in Rubisco activity was initially greater than the decrease in net photosynthesis (Hall et al., 1978). In a willow canopy, Rubisco-specific activity was higher when the apparent Et (N content in leaves) was smaller (Vapaavuori and Vuorinen, 1989). A similar nonlinearity was found in our previous experiments (Eichelmann and Laisk, 1990), in which we obtained a saturating relationship when Et exceeded 30 μmol m−2. In the latter work the initial slope of the A versus Cw curves under nonphotorespiratory conditions (1.5% O2) was assumed to represent the Rubisco activity in vivo and was compared with the Et. We discovered that growth light had the strongest influence on the saturation of the relationship between μ and Et. In the present work we present insight into this relationship, using not only plants grown under different light intensities but also leaves adapted to different light intensities. |
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