PPS '97 - Quaternary Structure: Multienzyme Complexes

Multienzyme Complexes

In a number of metabolic pathways, several enzymes which catalyze different stages of the process have been found to be associated noncovalently, giving a multienzyme complex. The proximity of the different types of enzymes increases the efficiency of the pathway: the overall reaction rate is increased with respect to catalysis by unassociated units, and side reactions are minimized. In some cases molecular mechanisms have been identified for the transfer of metabolites from one enzyme to the next within the complex. Later in the course we will be studying enzymes in general.

Pyruvate Dehydrogenase Complex

This multienzyme complex catalyses the conversion of pyruvate and coenzyme A (CoA) to acetyl CoA.

The reaction

There are four stages in this pathway, which are catalyzed by three enzymes:

"E1" - pyruvate dehydrogenase: This enzyme catalyzes the decarboxylation of pyruvate. This involves the prosthetic group thiamine pyrophosphate, or TPP.
"E2" - dihydrolipoyl transacetylase: Two steps of the pathway are catalyzed by this enzyme:
"E3" - dihydrolipoyl dehydrogenase: Finally, this enzyme regenerates the oxidized form of lipoamide. This involves the FAD prosthetic group.

Note that TPP, lipoamide and FAD are catalytic cofactors which remain unaltered by the net reaction, whereas CoA and NAD+ are stoichiometric cofactors; the overall reaction is:

pyruvate + CoA + NAD+ ----> acetyl CoA + carbon dioxide + NADH

The four stages are summarized in this diagram .

Note that the lipoamide cofactor of E2 reacts with the product (hydroxyethyl-TPP) from E1, and (in its modified form, dihydrolipoamide, formed by the third step) interacts with E3. It is for this reason that the interaction rate is increased by proximity of all three enzymes, and in fact the lipoamide group is a long flexible arm about 14Å long. It is covalently bonded to a specific lysine residue of the enzyme. The movement of the arm may be driven by a change in the net charge of the lipoyl group, depending on the ionization of the sulphydryl groups.

The structure of the complex

In isolation, E2 forms a homo-24-mer, believed to have cubic symmetry. The E1 and E3 subunits each form homodimers. Electron micrographs indicate that the whole complex has a cubic arrangment, in which an E3 dimer associates with each face of the E2 24-mer cube, while an E1 dimer is positioned on each edge of the cube.

Click here for a diagram of this model.

Refer to Reed(1974).

The Electron Transport Chain

Oxidative phosphorylation occurs in the mitochondria. The various components of the electron transport chain are situated in the inner membrane of this organelle. Oxidation of NADH and FADH2 (produced by the citric acid cycle) results in free energy which is coupled to the phosphorylation of ADP to form ATP (by ATP synthase, another complex in the inner mitochondrial membrane). Electrons pass from lower to higher standard reduction potentials in the chain in a series of redox reactions; the ultimate electron acceptor is oxygen (O2).

The electron transport chain in fact consists of four multienzyme complexes. These complexes are free to diffuse laterally through the membrane, and are not present in the same numbers.

Complex I: NADH - Coenzyme Q reductase, 26 subunits, 850 kD. This large protein includes one molecule of the prosthetic group flavin mononucleotide , or FMN, and six to seven iron-sulphur clusters
Complex II: succinate - Coenzyme Q reductase, 5 subunits, 127 kD. Includes:

FAD, covalently bound
three iron-sulphur clusters
cytochrome b560

Complex III: Coenzyme Q - cytochrome c reductase, 10 subunits, 280kD

cytochrome b562 256b (two molecules in asymmetric unit) (174Kb) [Bbk|BNL|ExP|Waw|Hal] SCRIPT
cytochrome b566
one iron-sulphur cluster
cytochrome c1

Complex IV: cytochrome c oxidase 7-8 subunits, 160-170kD

cytochrome a
cytochrome a3

Other components of the chain are succinate (between complexes I and II in the sequence), Coenzyme Q (between II and III) and

cytochrome c 1ccr (90Kb) [Bbk|BNL|ExP|Waw|Hal] SCRIPT (between III and IV).

Electron micrographs have indicated the overall shape of Complex IV and its orientation relative to the inner mitochondrial membrane. Techniques involving chemical cross-linking and antibody-binding to different subunits have led to a model of the whole complex, shown in this diagram (after Brunori and Wilson, 1982).The cytochrome c oxidase complex catalyzes the oxidation of four reduced cytochrome c molecules; the binding of cytochrome c to the complex mainly involves subunit II as indicated.

Refer also to Simon Brocklehurst's Multienzyme Complexes page.

References

Pyruvate Dehydrogenase Complex

Reed, L.J., (1974) Multienzyme complexes, Acc. Chem. Res. 7, 40-46
Stryer, L., (1981) Biochemistry, W.H. Freeman & Co., New York pp. 290-294
Voet, D. and Voet, J.G. (1990) Biochemistry, John Wiley & Sons, New York, pp. 186-188

Electron Transport Chain

Brunori, M. and Wilson, M.T. (1982) Cytochrome oxidase, Trends Biochem. Sci. 7, 295-299
Capaldi, R.A. (1982) Arrangement of proteins in the mitochondrial inner membrane, Biochim. Biophys. Acta 695, 291-306
Capaldi, R.A., Malatesta, F. and Darley-Usmar, V.M. (1983) Structure of cytochrome c oxidase, Biochim. Biophys. Acta 726, 135-148
Darnell, J., Lodish, H. and Baltimore, D. (1986) Molecular Cell Biology, W.H. Freeman & Co., New York pp. 873-889
Poulos, T.L. and Kraut, J. (1980) A hypothetical model of the cytochrome c peroxidase . cytochrome c electron transfer complex, J. Biol. Chem. 255 10322-10330
Scott, R.A. (1989) X-ray absorption spectroscopic investigations of cytochrome c oxidase structure and function, Annu. Rev. Biophys. Biophys. Chem. 18 137-158
Stryer, L., (1981) Biochemistry, W.H. Freeman & Co., New York pp. 319-320
Sweeney, W.V. and Rabinowitz, J.C. (1980) Proteins containing 4Fe-4S clusters: an overview, Annu. Rev. Biochem. 49, 139-161
Voet, D. and Voet, J.G. (1990) Biochemistry, John Wiley & Sons, New York, pp. 532-544

Section 11 Index

Index to Course Material

Multimeric Enzymes

Last updated 14th April '97