Interaction of polyethylene glycol-6000 with C4 phosphoenolpyruvate carboxylase in crude leaf extracts as well as in purified protein form from Amaranthus hypochondriacus L.: evidence for oligomerization of PEPC in vitro and in vivo.

The interaction of phosphoenolpyruvate carboxylase (PEPC) with a compatible solute, PEG-6000, was examined using crude leaf extracts as well as the purified protein from leaves of Amaranthus hypochondriacus, a NAD-malic enzyme type C4 plant. The inclusion in the assay medium of PEG-6000 stimulated the activity of purified PEPC by about 2.5-fold over control. The addition of PEG during both extraction and assay, stimulated PEPC activity by almost 5.0 fold in crude extracts. The stimulation by PEG of the dark-form of PEPC (2.4 fold) was more than that of the light-form (1.7 fold). Gel filtration of PEPC in leaf extracts on Sephadex G-200, showed the existence of three different oligomeric forms: tetramer, dimer and monomer. The exclusion of PEG and glycerol during extraction and elution on Sephadex resulted in a marked shift of the enzyme into dimer and/or monomer, with a very small proportion of tetramer but on the contrary, the inclusion of PEG and glycerol resulted in the enzyme maintaining predominantly a tetrameric shape. Thus, the activity and the structural properties of PEPC can be influenced by the presence or absence of compatible solutes (PEG or glycerol), obviously due to changes in the microenvironment of the enzyme.