Phosphoenolpyruvate Carboxylase During Development of Crassulacean Acid Metabolism and During a Diurnal Cycle inMesembryanthemum Crystallinum

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Abstract
Crassulacean acid metabolism (CAM) in Mesembryanthemum crystallinum was induced by transfer of plants from 100 to 400 mM NaCl. Diurnal malate fluctuations developed slowly; maximum rates of net malate synthesis in the dark were reached only on the 10th day after NaCl was increased to 400 mM. In contrast, phosphoenolpyruvate carboxylase (PEPC) activity, assayed at optimum pH of 8–0, had nearly reached its maximum on the 5th day after plants were transferred to 400 mM NaCl. Characteristics of PEPC changed during the first 12 d of exposure of plants to 400 mM NaCl. There were increases in the ratio of PEPC activity at pH 7 0/PEPC activity at pH 8.0, and decreases in the Km for PEP measured at pH 7.0, and possibly in the degree of malate inhibition. All further measurements were made once CAM was well established. In vivo rates of malate synthesis were 14–18 times smaller than PEPC activity at 2 mM PEP, both processes being measured at 15 °C. It is suggested that the high PEPC levels favour rapid, preferential flow of carbon to malate, by maintaining very low PEP levels in the cytoplasm. PEPC changed in characteristics during the diurnal cycle. During the first few minutes after isolation, extracts made during the first hours of the day, when malate was consumed, showed very low PEPC activity at pH 7.0 but high activity at pH 8.0. The activity of PEPC at pH 7.0 rose gradually during storage of the extracts at 0 °C, usually reaching the activity at pH 8.0 after about 30–50 min. In contrast, extracts obtained during the first hours of the night, when malate was synthesized, showed high PEPC activity at both pH 7.0 and 8–0 within 30–50 s after extraction. The results indicate that PEPC of M. crystallinum, performing distinct CAM, may exist in two states. One state would favour rapid malate synthesis and transport to the vacuoles and would function during the night. The second state, with little activity below pH 7.5, would occur during the day, thus preventing complications of continued synthesis of malate while it is converted to carbohydrates.