Hyperactive enzymes through complex formation with customized polymers.
The use of enzymes as a substitute for chemical catalysts is becoming increasingly important in synthetic chemistry. In this context, boosting enzyme activities beyond the level of their natural activities is of particular interest. Various strategies are currently being pursued to increase enzyme activity, ranging from engineering the active site and immobilizing the enzyme to optimizing the reaction environment or adding further substances. Various polymers are currently being discussed as suitable additives that interact with the surfaces of the enzymes and can significantly increase their natural activity.
Scientists of Potsdam University together with the IPB's computer modeling experts have now tested the hyperactivation of the endopeptidase α-chymotrypsin through the addition of acrylic acid-based polymers. In these assays, they tested not only the poly(acrylic acids) (PAA) that are usually used to increase enzyme activity, but also self-produced synthetic compounds, of which poly(2-carboxyethyl) acrylate (PCEA) proved to be particularly effective. Activity assays of α-chymotrypsin with N-glycyl-L-phenylalanine-p-nitroanilide (GPNA) as the substrate to be cleaved showed enormous increases in the endopeptidase activity of 950% when PCEA was added and 450% when PAA was used as an additive.
The mechanism of this hyperactivation is not yet completely clear, but the IPB computational chemists were able to shed more light on it. In a combined experimental and computer-aided analysis, the scientists investigated the interactions between α-chymotrypsin and the polymers. Using isothermal titration calorimetry, they were able to demonstrate a pronounced complex formation between the polymer and the enzyme. Interestingly, docking studies revealed a significantly increased binding affinity of the substrate to the enzyme-polymer complex compared to the enzyme alone. This increased binding affinity could be due to an enlarged active site of the enzyme in the presence of the polymers - which was shown by molecular dynamics simulations. The enlargement of the active site cavity may improve substrate access, which in turn may lead to the enormous increase in activity of the enzyme-polymer complexes. The present study provides important insights into the molecular mechanisms of enzyme hyperactivity, the scientists conclude. The knowledge gained forms the basis for developing further synthetic polymers that will be used to optimize biocatalytic syntheses.
