PEPC structure

Structure of PEPC

From Escherichia Coli

Phosphoenolpyruvate Carboxylase functions as a homotetramer. Figure 2 from fig. 1 of Kai et.al (1999) is the PEPC homotetramer. The PEPC homotetramer has four identical subunits each represented as a different color in Fig. 2. The molecular masses of each subunit is 95-110kDa, and each subunit contains its own catalytic and allosteric site. Fig. 2 (a) Ribbon diagram showing the PEPC homotetramer. (b) Ribbon diagram (same as a) rotated 90 degrees. (c and d) a spacefill model of PEPC. The orientations are the same as in a and b. The spacefill model (c) shows a close interaction between the lavender and blue subunits and between the pink and green subunits. These close interactions show the arrangement of PEPC in a"dimer-of-dimers". The dimers are held together through an interation between Arg -438 of one subunit and Gly-433 of the neighboring subunit, forming a salt bridge (Kai et. al., 1999). These salt bridges are disrupted when the tetrameric enzyme is dissociated into two dimers.

Fig. 2

MONOMER of PEPC

Each subunit of PEPC contains the following:

(Push buttons in the order they come to see the full affect)

8 beta-strands forming a single beta barrel (click button to view)

40 alpa helices (click button to view)

L-aspartate and L-malate are allosteric inhibitors of PEPC in E.coli. L-malate is an allosteric inhibitor of C₄ plants.

L-Aspartate is one of the allosteric effector molecules for PEPC that causes inhibition of the catalytic activity (Kai, 1999). The binding domain for the allosteric inhibitor consists of four amino acid residues, Lys-773, Arg-832, Arg-587, and Asn-881. All four of these residues are highly conserved in all PEPCs so the information stated just doesn't hold true for E. coli. Lys-773 and Arg-832 forms a salt-bridge with the carboxyl group in the side chain of aspartate. Arg-587 has 6 glycine residues surrounding it that provide a long flexible loop which is locates the allosteric site near the enzymes active site. The Arg-587 also forms a salt-bridge to the carboxyl group of aspartate, but unlike the Lys and Arg-832, Arg-587 is trapped by the aspartate. The Asn-881 forms hydrogen bonds with the aspartate.

Fig. 3 (PDB file 1FIY)

(Click here to view L-aspartate)

(Click here to view the allosteric binding domain)

Lys-773= red; Arg-832=blue; Arg-587=green; Asn-881=purple

Active Site

The active site is located at the C-terminus of the beta barel. Two highly conserved histidine residues are located in this area and are important for the catalytic activity. Histidine-138 and Histidine-579 can be seen in Fig. 4, from Fig. 6 by Kai, et. al., 1999, along with another important residue Arg-587. Arg-587 is involved in the allosteric binding of L-Aspartate. The interatomic distances between the alpha-carbon atoms of the histidines and the aspartate range from 17 angstroms to 18 angstroms (Kai et. al., 1999).

Fig. 4 (Stereoview of the active site of PEPC)

The residues involved in the catalytic active are shown in ball- and- stick representation. Can you identify the important residues His-138, His-579, and Arg-587? The missing loop is shown as dots. The beta sheets of the beta barel are shown in green, while the yellow and red regions are the alpha helices. The residues are all located at the C-termini. The missing loop is missing Lys-702 to Gly-708. It is here that the active site is located. Lys-702 and Arg-704 are highly conserved in all PEPCs identified. The missing loop, consisting of Gly-707 and Gly-708, trap essential functional residues with the help of Ala-701 or Ser-701. The link between the catalytic site and the allosteric binding domain is the glycine loop and the loop consisting Arg-587 and Arg-581. These two loops work together to catch substrate molecules at the active site with the their functional residues; plus they form lids protecting the reaction intermediates from outside water molecules (Kai et.al., 1999). The C-terminus of the beta barel contains 19 residues and 14 of these residues are hydrophobic making them essential for keeping water away from the reaction.

Click here to view residues His-138, His-579, and Arg-587.

His-138=red; His-579=blue; Arg-587=green

The missing loop is not present in the PDB of PEPC, hence the reason for the dot line representation. The exact catalytic site cannot be viewed.

Click here to see how close the active site is to the allosteric site with the binding of L-aspartate.

Click here to view the allosteric site, the active site and L-aspartate (The colors used are the same as before. The last two molecules to change to red and blue are His-138 and His-579, respectively.)

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