Phosphoenolpyruvate Carboxykinase in C4 Plants: Its Role and Regulation

Abstract

Some of the recent findings which revise our view of the role and regulation of phosphoenolpyruvate carboxykinase (PEPCK) in C4 plants are discussed. Evidence is presented that PEPCK is present at appreciable activities in the bundle-sheath of some NADP-malic enzyme-type C4 plants, such as maize, but it was not detectable in NAD-malic enzyme-type C4 plants. PEPCK is rapidly inactivated in crude extracts of leaves of the C4 plant, Panicum maximum. This inactivation could be prevented by high concentrations of dithiothreitol or by the inclusion of ADP or ATP, suggesting the involvement of thiols at the active site. PEPCK is also subject to rapid proteolysis in crude extracts of a range of C4 plants, resulting in cleavage to a smaller (62 kDa) form. This can be reduced by extraction at high pH and by the inclusion of SDS, but it means that intact PEPCK has never been purified from a C4 plant. The molecular mass of PEPCK varies considerably in C4 plants, unlike C3 and CAM plants in which it is usually 74 kDa. PEPCK is phosphorylated during darkness (and reversed by light) in some C4 plants with PEPCK of a larger molecular mass, such as Panicum maximum (71 kDa), but it was not phosphorylated in the PEPCK-type C4 plant, Sporobolus pyramidalis (69 kDa). The known regulatory properties of PEPCK are discussed in relation to its role in C4 photosynthesis, in particular its sensitivity to regulation by adenylates and by Mn2+.

Phosphoenolpyruvate Carboxykinase Is Involved in the Decarboxylation of Aspartate in the Bundle Sheath of Maize1

“…1) and activities (Table I) of PEPCK. The activity of PEPCK recorded in maize was between one-fifth and one-half of that measured in PEPCK C 4 plants (Chapman and Hatch, 1983;Smith and Woolhouse, 1983;Walker et al, 1997).…”

“…Even though this treatment did not lead to general proteolysis, it resulted in the cleavage of PEPCK. PEPCK in plant extracts is rapidly cleaved at low pH or, in the absence of SDS, to a 62-kD form with no loss of activity (Walker et al, , 1997. However, during the isolation of BS strands from maize, PEPCK was degraded further and sometimes no PEPCK protein was detectable on western blots.…”

“…Other NADP-ME species such as Echinochloa colona, Echinochloa crus-galli, Digitaria sanguinalis, and Paspalum notatum also contain PEPCK, whereas PEPCK protein was not detectable in Sorghum bicolor, Saccharum officinarum, or Flaveria bidentis (Walker et al, 1997). The reason that the occurrence of PEPCK in these plants has been overlooked may be due to difficulties in measuring PEPCK activity (Walker et al, 1997) or to the lack of an antibody.PEPCK-type C 4 species mainly use Asp as the CO 2 donor and decarboxylate the oxaloacetate formed via the following reaction:In NADP-ME species such as maize, 14 CO 2 is initially incorporated into the C-4 position of malate and Asp (about 75% and 25%, respectively), and the C-4 of both is subsequently incorporated into other metabolites (Hatch, 1971; Morot-Gaudry and Farineau, 1978). Using isolated BS strands, Chapman and Hatch (1981) showed that BS cells of maize have a significant capacity to decarboxylate Asp, and they assumed that NADP-ME was responsible.…”

“…Maize (Zea mays) belongs to the NADP-ME subgroup of C 4 plants and possesses negligible activity of PEPCK, although suggested that some other NADP-ME species utilize PEPCK as an auxiliary decarboxylase. However, we showed previously that maize leaves contain large amounts of PEPCK (Walker et al, 1997), andFurumoto et al (1999) identified a PEPCK gene from maize that is specifically expressed in BS cells. Other NADP-ME species such as Echinochloa colona, Echinochloa crus-galli, Digitaria sanguinalis, and Paspalum notatum also contain PEPCK, whereas PEPCK protein was not detectable in Sorghum bicolor, Saccharum officinarum, or Flaveria bidentis (Walker et al, 1997).…”

“…However, we showed previously that maize leaves contain large amounts of PEPCK (Walker et al, 1997), andFurumoto et al (1999) identified a PEPCK gene from maize that is specifically expressed in BS cells. Other NADP-ME species such as Echinochloa colona, Echinochloa crus-galli, Digitaria sanguinalis, and Paspalum notatum also contain PEPCK, whereas PEPCK protein was not detectable in Sorghum bicolor, Saccharum officinarum, or Flaveria bidentis (Walker et al, 1997). The reason that the occurrence of PEPCK in these plants has been overlooked may be due to difficulties in measuring PEPCK activity (Walker et al, 1997) or to the lack of an antibody.…”