Chemists make important discovery on how enzymes work

University at Buffalo chemists report the discovery of a central mechanism responsible for the action of the powerful biological catalysts known as enzymes. They published their results in an open access article in the journal Biochemistry. The findings surprised many enzymologists.

The UB research provides critical insight into why catalysis is so complex and may help pave the way for improving the design of synthetic catalysts. Such catalysts are expected to be used widely in the production of cellulosic biofuels and in biorefining. In one such example, showing what the future may hold, scientists recently designed synthetic enzymes from scratch and found they were highly efficient in the catalytic conversion of starch and sugar (with water) into biohydrogen (previous post). The new discovery about the essence of enzymatic catalysis may spur the development of similar applications.

Enzymes are the products of billions of years of cellular evolution. Attempts to replicate evolution and design catalysts of non-biological reactions with enzyme-like activity have failed, because scientists have yet to unravel the secrets of enzyme catalysis. The more that is known about catalysis, the better chances we have of designing active catalysts. - John P. Richard, Ph.D., co-author and professor of chemistry at the UB College of Arts and Sciences

Together with Tina L. Amyes, Ph.D., UB adjunct associate professor of chemistry, Richard thinks the discovery will have the potential to transform the chemical industry in processes ranging from soft-drink manufacturing to the production of ethanol and countless other industrial processes.

While attempts to design catalysts have been somewhat successful, the catalysis that results is far less efficient than that produced by reactions with enzymes.

Non-reactive substrate portion key
Protein catalysts are distinguished by their enormous molecular weights, ranging from 10,000 to greater than 1,000,000 Daltons, whereas a synthetic molecule with a weight of 1,000 would be considered large. The recent results by Richard and Amyes provide critical insight into why effective catalysis requires such large molecules. Catalysis starts with molecular recognition of the substrate by the catalyst.

The so-called "catalytic" recognition is limited in man-made catalysts to several atoms that participate in the chemical reaction. Amyes and Richard have provided compelling evidence that interactions between enzymes and non-reacting portions of the substrate are critical for large catalytic rate accelerations:
bioenergy :: biofuels :: energy :: sustainability :: biomass :: bioconversion :: biorefining :: enzyme :: catalysis ::

These findings demonstrate a simple principle of catalysis that is important for many enzymes that catalyze reactions of substrates containing phosphate groups and which can be generalized to all enzymes.

The chemistry between a catalyst and substrate occurs where groups of amino acid residues interact with the substrate. But enzymes also have domains that interact with the non-reacting parts of the substrate.

A flexible loop on the enzyme wraps around the substrate, burying it in an environment that is favorable for catalysis. In order to bury the substrate, certain interactions are necessary that allow the loop to wrap around the substrate and that is what the phosphate groups on the substrate are doing.

The UB research demonstrates just how important this process is to catalysis. Richard and Amyes discovered these interactions are critical to the process of making reactions faster.

Experimental method
The critical experiment by the UB researchers was to clip the covalent bond that links the phosphate groups to the substrate. "We have found that the interactions between phosphate groups and several enzymes are used to promote the chemistry even in the absence of a covalent linkage," said Richard. "These results have surprised many enzymologists."

To conduct the research, Richard and Amyes developed a specialized and technically difficult assay for enzyme activity that uses nuclear magnetic resonance spectroscopy to detect chemical reactions that would normally be invisible.

Image: Nature breaks and forms the strongest chemical bonds with incomparable efficiency using enzymatic catalysis. In living cells enzymes catalyze, for instance, the synthesis of proteins and DNA, the cleavage of carbohydrates and proteins and the transformation of toxic side products of the respiration cycle into harmless compounds. In each case the chemical transformation occurs with high selectivity and at an exceptionally high rate under physiological conditions. The major source of the catalytic power of enzymes is the stabilization of the transition state relative to the reactant and in certain cases and to a smaller extent an increase of tunneling effects. The combined catalytic effects lead to rate enhancements of up to 1019 relative to the uncatalyzed reaction in solution. The image shows the structure of an enzyme, endoprotease thermolysin. The active site is depicted in stick and ball representation. Thermolysin catalyzes the cleavage of peptide bonds by 5-7 orders of magnitude relative to alkaline hydrolysis in aqueous solution.

More information:
Tina L. Amyes and John P. Richard, "Enzymatic Catalysis of Proton Transfer at Carbon: Activation of Triosephosphate Isomerase by Phosphite Dianion", Biochemistry, 2007; 46(19) pp 5841 - 5854; (Article) DOI: 10.1021/bi700409b

University at Buffalo: How Enzymes Work: UB Chemists Publish A Major Discovery - June 20, 2007.