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bioRxiv · 10.1101/2023.05.02.539073

Enzyme stability-activity trade-off: new insights from protein stability weaknesses and evolutionary conservation

Hou, Q.·Rooman, M.·Pucci, F.
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Abstract

A general limitation of the use of enzymes in biotechnological processes under sometimes non-physiological conditions is the complex interplay between two key quantities, enzyme activity and stability, where the increase of one is often associated with the decrease of the other. A precise stability-activity trade-off is necessary for the enzymes to be fully functional, but its weight in different protein regions and its dependence on environmental conditions is not yet elucidated. To advance this issue, we used the formalism that we have recently developed to effectively identify stability strength and weakness regions in protein structures, and applied it to a large set of globular enzymes with known experimental structure and catalytic sites. Our analysis showed a striking oscillatory pattern of free energy compensation centered on the catalytic region. Indeed, catalytic residues are usually non-optimal with respect to stability, but residues in the first shell around the catalytic site are, on the average, stability strengths and thus compensate for this lack of stability; residues in the second shell are weaker again, and so on. This trend is consistent across all enzyme families. It is accompanied by a similar, but less pronounced, pattern of residue conservation across evolution. In addition, we analyzed cold- and heat-adapted enzymes separately and highlighted different patterns of stability strengths and weaknesses, which provide insight into the longstanding problem of catalytic rate enhancement in cold environments. The successful comparison of our stability and conservation results with experimental fitness data, obtained by deep mutagenesis scanning, led us to propose criteria for improving catalytic activity while maintaining enzyme stability, a key goal in enzyme design.

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Hou, Q., Rooman, M., Pucci, F.. 2023-05-03. Enzyme stability-activity trade-off: new insights from protein stability weaknesses and evolutionary conservation. https://doi.org/10.1101/2023.05.02.539073

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