Electron bifurcation: a new mechanism of energy coupling

 

Consider a reaction: A -> B that is endergonic. Without the input of energy, the reaction would not proceed. However, if we would couple the reaction to a second:

 

A + B -> C + D

 

it could proceed if C -> D could provide the energy for A ->B. Such energetic coupling reactions are widespread in biology and found in many biochemical reactions. So far, two driving forces have been known: one is the hydrolysis of ATP (or other nucleoside triphosphates) and the other the electrochemical ion potential across the cytoplasmic membrane. Whereas the former is generally used in biochemical pathways (for example: Glucose + ATP -> Glucose-6-P + ADP), the latter is used to drive redox reactions (for example, uphill transport of electrons from an electropositive donor (Fe2+ -> Fe3+) to a more electronegative acceptor (NAD -> NADH2). Another example is NADH-dependent reduction of ferredoxin, catalyzed by the Rnf complex.

 

In 2008, a fundamentally different mechanism of driving endergonic redox reactions was discovered by Buckel and Thauer. They found a soluble enzyme that couples an endergonic reaction A ->  B to an exergonic reaction A -> C. They discovered that the butyryl-CoA dehydrogenase of Clostridium kluyveri drives endergonic electron flow from NADH (E0’ = -320 mV) to ferredoxin (E0’ ~ -500 mV) by a simultaneous electron flow from NADH to crotonyl-CoA (thus producing butyryl-CoA). The enzyme (butyryl-CoA dehydrogenase) is organized in a complex with an electron-transfer flavoprotein (Etf) and the flavin is seen as the site where the electrons are bifurcated to the two acceptors with different redox potentials.

 

The mechanistic basis of this novel and eminent important coupling mechanism (since it produces reduced ferredoxin, the “fuel” of the respiratory enzymes in these anaerobes, see topic III) is far from being understood.

 

Our group has contributed a number of novel electron bifurcating enzymes and we are currently in the process of trying to understand how they function.

 

 

1. The electron bifurcating hydrogenase of A. woodii

 

A. woodii grows on H2 + CO2 and is challenged by the task to reduce ferredoxin (E0’ ≈ - 500mV) with H2 (E0’ = - 414 mV). The solution to this problem is electron bifurcation.  Hydrogen is oxidized and the electrons are bifurcated to ferredoxin and NAD. The reaction catalyzed by the enzymes is reversible. Thus, this enzyme is a key component in anaerobic metabolism and in biological hydrogen production.

Model of the electron-bifurcating hydrogenase of A. woodii. Fd, ferredoxin,

 

References

Schuchmann, K., Müller, V. (2012) A bacterial electron bifurcating hydrogenase. J. Biol. Chem. 287 : 31165-31171.

 

 

 

2. The electron bifurcating caffeyl-CoA reductase of A. woodii

 

This enzyme plays an essential role in caffeate respiration (see topic II) It couples exergonic electron flow from NADH to caffeyl-CoA with the endergonic reduction of ferredoxin (with NADH) as reductant. Reduced ferredoxin fuels the Rnf complex, the only energy-conserving respiratory enzyme of this respiration. The enzyme is composed of the reductase and the Etf protein. It contains flavins and iron sulfur centers. The structure-function analysis of this enzyme is currently under way in our laboratory.

 

Model of the electron-bifurcating caffeyl-CoA reductase/electron transfer flavoprotein complex. Etf, electron transfer flavoprotein; Car, caffeate respiration; Fd, ferredoxin.

 

 

References

Bertsch, J., Parthasarathy, A., Buckel, W., Müller, V. (2013) An electron-bifurcating caffeyl-CoA reductase. J. Biol. Chem. 288 : 11304-11311.

Hess, V., González, J.M., Parthasarathy, A., Buckel, W., Müller, V. (2013) Caffeate respiration in the acetogenic bacterium Acetobaceterium woodii: a CoA loop saves energy for caffeate activation. Appl. Environ. Microbiol. 79 : 1942-1947.

Hess, V., Vitt, S., Müller, V. (2011) A caffeyl-coenzyme A synthetase initiates caffeate activation prior to caffeate reduction in the acetogenic bacterium Acetobacterium woodii. J. Bacteriol. 193 : 971-978.

 

 

 

3. The electron confurcating lactate dehydrogenase of A. woodii

 

This enzyme is essential in lactate metabolism in A. woodii (see topic II). Oxidation of lactate with concomitant reduction of NAD is endergonic and driven by the simultaneous exergonic electron flow from reduced ferredoxin to NAD according to:

 

lactate + Fd2- + 2 NAD   ->   pyruvate + Fd + 2 NADH

 

Again, the reaction is fully reversible.

 

The lactate dehydrogenase is in a complex with an Etf protein (different from the one in the caffeyl-CoA reductase). It contains flavins and iron sulfur centers and its function is currently being analyzed in our lab.

 

Future work is aimed to produce these enzymes from A. woodii heterologously or homologously. This is the first step to a molecular analysis based on site-directed mutagenesis.

Model of the electron-confurcating lactate dehydrogenase/electron transfer flavoprotein complex. Etf: electron transfer flavoprotein; Fd: ferredoxin.

 

 

References

 

Weghoff, M.C., Bertsch, J., Müller, V. (2014) A novel mode of lactate metabolism in strictly anaerobic bacteria. Environ. Microbiol., doi:10.1111/1462-2920.12493.

 

Highlight in:
Schink, B. (2014) Electron confurcation in anaerobic lactate oxidation. Environ. Microbiol., in press.