Análise da variação de estabilidade conformacional de proteínas decorrente de mutações pontuais

Abstract

Proteins have a number of biotechnological applications and are particularly convenient for their specificity. However, the industrial use of proteins can be limited by their lack of stability in high temperatures, insufficient affinity by the substrate and the number of reactions it catalyzes, whereas the therapeutic use is restricted by their potential immunogenicity. A protein can be engineered for new purposes, such as increased affinity, via a series of mutations in its primary sequence, but all protein engineering projects must account for the protein stability. Site-directed mutagenesis is the cheapest way of generating new mutants, however, in silico methods are still inefficient in predicting how it should be preferrably conducted. This work aimed to study how mutation site and amino acid features reflect on the change of stability of a protein structure that undergoes a single-point mutation. The investigation was based on an experimental dataset of 3605 single-point mutants along with their DDG values, which were used to classify mutations in two groups of interest: neutral mutations, when there is no significant effect on stability, or favorable ones, when DDG is smaller than - 1 kcal/mol. Two structural features were studied to account for choosing the most suitable mutation site for each group of interest: solvent accessibility surface area (SASA) and secondary structure (SS); and three amino acid features were studied to investigate how amino acid identity can influenciate on protein stability changes: volume, molar mass and hydrophobicity. Results show that SASA values influenciate directly on stability, since SASA values can suggest if the mutation site is near the hydrophobic core. Secondary structure, however, does not play a significant role on separating neutral mutations from favorable ones according to the results. Molar mass and volume are found to be equivalent and important to stability, however, results on hydrophobicity suggest a low relevance of this feature on protein stability changes. The results are elucidative but further analysis of these descriptors, as well as investigation of new ones, are still necessary to accomplish mutation features prediction.