How to Make an Enzyme Accept a Smaller Substrate

Manfred Reetz and his team have published an example of a method that may prove to be a general way to persuade an enzyme normally acting on a large substrate to accept a much smaller one. What is the secret? By partially filling the binding pocket with a non-reactive guest compound that also has affinity for the desired smaller compound, affinity for the desired small target compound is increased and catalysis occurs. Importantly, the non-reactive guest is selected so that space remains to accommodate the smaller target compound in the area of the binding pocket.

Using a cytochrome P450 enzyme that normally acts on a long-chain fatty acid as its preferred substrate for oxidation, Reetz and co-workers found that by giving the enzyme an appropriately-sized perfluorocarboxylic acid to partially fill the binding pocket, a smaller hydrocarbon substrate could be accommodated as well in the same pocket. In this way, the Cytochrome P450, which catalyzed hydroxylation of a fatty acid, becomes a methane mono-oxygenase.

This development is a step in the direction of the biocatalytic conversion of methane into methanol, but there may be broader implications if this strategy can be generalized to other enzymes. Using the guest strategy in combination with directed evolution will open up many new possibilities for enzyme engineering.