Activation of Rubisco regulates photosynthesis at high temperature and CO2.

T enzyme Rubisco, short for ribulose1,5-bisphosphate carboxylaseyoxygenase, is the enzyme that incorporates CO2 into plants during photosynthesis. As it constitutes about 30% of the total protein in a plant leaf, Rubisco is probably the most abundant protein on earth and a major sink for plant nitrogen. Rubisco is widely accepted as the ultimate rate-limiting step in photosynthetic carbon fixation. Atmospheric oxygen competes with CO2 as a substrate for Rubisco, giving rise to photorespiration. When first purified, the enzyme appeared to have a poor affinity for CO2 [Km (CO2)] of 450 mM (1), whereas air in equilibrium with water, 25°C, is about 10 mM. Later, Lorimer et al. (2) showed that the active site of Rubisco must first be carbamylated by an activator CO2, separate from the substrate CO2, and must bind Mg21 before binding the five-carbon substrate, ribulose-1,5-bisphosphate (RuBP). Indeed, on addition of RuBP, the measured Km(CO2) approached the concentrations of dissolved CO2 in water; however, the rate of the reaction was sustained for only 5 minutes and then declined rapidly. This decline was shown to be caused by the tight binding of RuBP to Rubisco that had lost the activator CO2 (Fig. 1). The missing ingredient needed to uncouple Rubisco and RuBP was found in the intact plant with a separate protein, called Rubisco activase (3). This enzyme acts on Rubisco and allows release of the bound RuBP so that the site can bind the activator CO2 and Mg21. Rubisco activase itself requires ATP, and its activity is related to the energy charge of the chloroplast (4). Thus, the proportion of Rubisco that is active in a leaf (activation state) can vary depending on the effectiveness of Rubisco activase in removing bound RuBP. Regulation of Rubisco fine tunes the rate of CO2 fixation to the rate of photosynthetic electron transport, ensuring that chloroplast metabolites are always optimal for photosynthesis (5). The paper by Crafts-Brandner and Salvucci (6) in this issue of PNAS provides evidence that, with plants under heat stress, the activation state of Rubisco and photosynthesis as measured by CO2 exchange is reduced. By duplicating the temperature response in the test tube under controlled conditions, and by using Rubisco and Rubisco activase isolated from tobacco, they were able to ascribe the limitation to a specific biochemical event, the inability of Rubisco activase to keep pace with a faster deactivation of Rubisco. Increased CO2 also decreased the activation state of Rubisco in leaves, and the authors conclude that the response could be explained by a decreased energy charge in the chloroplast that reduced the ATPase activity of Rubisco activase. By calculating photosynthesis on the basis of the kinetics of Rubisco and the amount of active enzyme in the leaf, the authors have shown that, under both high temperature and high CO2, photosynthesis was constrained by the activity of Rubisco activase. Changes in the global environment since the beginning of the Industrial Revolution have raised concerns about the impact on natural and agroecosystems. Increasing CO2 levels and corresponding