In Vivo Properties of Phosphoenolpyruvate Carboxylase in Crassulacean Acid Metabolism Plants. Is Pineapple CAM Not Regulated by PEPC Phosphorylation?

Abstract

The in vivo sensitivity to inhibition by L-malate of phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31) in rapidly prepared crude extracts (within 2 minutes) was investigated using two CAM plants, Kalanchoe pinnata and pineapple (Ananas comosus cv. Smooth-cayenne N67-10). The phosphorylation process may not occur in pineapple PEPC. Variations were also observed in apparent kinetic properties and molecular mass as well as in the subunit composition of PEPC from these plants. Two different subunits (major polypeptide 112 kDa and minor polypeptide 119 kDa) were separated from day and night forms of PEPC from K. pinnata; only one identical subunit (107 kDa) was separated from pineapple. The integrity of both forms of in vivo PEPC from these plants was demonstrated by Western blot analyses using anti-N-terminus antibody. K. pinnata lost approximately 87% of malate [malic acid] sensitivity, whereas pineapple PEPC lost only 21% sensitivity at night in the presence of 2 mM L-malate. The apparent Ki (L-malate) was 6.4- fold higher in phase I (5.8 mM) than in phase III (0.9 mM) from K. pinnata; however, it was only 1.0 and 0.5 mM for day and night PEPC, respectively, from pineapple. These findings suggest that the phosphorylation process may not be the major regulatory mechanism in pineapple PEPC.