INSIGHTS INTO STRUCTURAL STABILITY DETERMINANTS OF NUCLEOSIDE HYDROLASES

"Thermophilic enzymes face the challenge of maintaining optimal catalytic activity at temperatures nearing the boiling point of water, hence, they can be used as interesting model systems to understand the evolution of enzymes, as well as the molecular strategies developed as defence against thermal denaturation. Several comparisons between the structures of homologous isozymes from mesophilic and thermophilic organisms highlighted specific properties of folded proteins that may contribute to thermal stability. Moreover, the understanding of general strategies that may enhance the stability of enzymes at high temperatures could be beneficial for the engineering of biocatalysts in industrial processes. . Here we report a comparative structure-thermostability analysis of a panel of tetrameric nucleoside hydrolase (NH) enzymes from mesophiles and thermophiles by a combined structural and spectroscopic characterization that utilize principal component analysis (PCA) and differential scanning fluorimetry (DSF). . The availability of high-resolution structures of NH enzymes isolated from both mesophilic and hyperthermophilic organisms belonging to the same structural homology group, emphasizes the elements that lead to enhanced stability and underscores that resistance to thermal denaturation can be achieved using a combination of different structural solutions. Apparently, each organism can employ a different combination of stabilizing structural features to achieve the required level of NH stability. However, the multivariate analysis highlights how the NHs from mesophilic and thermophilic organisms here considered can be clustered according to the type and extent of specific features used for optimization. The results obtained indicate that the inclusion of salt bridges at the intersubunit contact surfaces is the only shared strategy in both mesophilic and thermophilic organisms to modulate the stability of NHs.. "