An in silico analysis of laccase thermostability determinants for the improvement of rational enzyme engineering
Welcome to project website for this bachelor’s thesis!
Laccases are enzymes that are widely distributed in nature, capable of degrading a wide range of substrates. Thanks to this property, they could be incredibly useful in bioremediation.
What is bioremediation?
Bioremediation refers to the treatment of different contaminants through the use of microorganisms or the enzymes they produce, capable of degrading these pollutants into less harmful compounds.
Unfortunately, many laccases are not very heat-resistant, which restricts their practical application. For this reason, this project set out to find what differentiates laccases with this property on a structural and physicochemical level.
To achieve this, thermostable and non-thermostable laccases were selected from different databases, aligned, and compared using statistical tests and the Kullback-Leibler divergence.
What is the Kullback-Leibler divergence?
The Kullback-Leibler divergence is a mathematical measure of how different two probability distributions are from one another. In the context of this project, it was used to compare the frequency of each of the 20 amino acids at equivalent positions between thermostable and non-thermostable laccases. A higher Kullback-Leibler divergence value at a given position indicates that the two groups of laccases tend to use different amino acids there, making it a useful tool for identifying the positions that differ most between thermostable and non-thermostable enzymes.
Through these tests, the project uncovered that thermostable and non-thermostable laccases are really quite similar on the whole, but variations in how they use specific amino acids at given positions reveal that thermostability may be determined by a small number of key locations in the protein structure rather than by widespread differences in composition. Many of these positions are found in flexible regions of the protein, such as loops connecting more rigid structural elements, suggesting that thermostable laccases may achieve their stability by locking down regions that would otherwise move too freely at high temperatures.
Identifying these positions is the first step toward engineering laccases that are more thermostable and, therefore, more useful in real-world bioremediation applications.