BindScan computer assisted protein design. Estudi del cas concret de la reacció de transglicosilació de la Lacto-N-Biosidasa


Romero Simo, Marc


In breast milk, HMOs (Human Milk Oligosaccharides) are very abundant and have a great importance since they are not found in any other type of milk. Despite not being capable of degrading them due to the lack of specific enzymes, they are a fundamental component in the feeding of new-borns since they act as prebiotics and allow in the intestinal track of new-borns, the development of microorganisms that provide great benefits by stimulating and protecting the immune system. An example of these microorganisms is Bifidobacterium Bifidum which uses Lacto-N-Biosidase to degrade HMOs through a hydrolysis reaction giving lactose and Lacto-n-Biose.
There is currently a line of research in this regard orientated into the synthesis of these oligosaccharides in order to supplement infant milk formula. To synthesize the HMO, using Lacto-N-Biosidase as protein and lactose as ligand, it is sought maximizing the reverse hydrolysis reaction, that is, transglycosylation. In order to enhance this reaction that occurs very rarely in nature, they are sought, thanks to the bioinformatics, possible mutations in Lacto-N-Biosidase that promote binding with lactose and therefore increase the efficiency of transglycosylation.
For this project, we have started from results previously found in the group of biochemistry to continue the research. First, a cluster analysis has been done to visualize the different conformations of lactose bound to Lacto-n-biosidase. Subsequently, by means of Bindscan, a new conformation of lactose has been explored in Lacto-N-Biosidase binding with the fixed W394F mutation, thus searching for possible mutants doubles that increase the binding energy and therefore promote transglycosylation.
In parallel, a new metric (Low RMSD Range) has been implemented in the algorithm in order to avoid considering false positive results. In this case it only shows mutants with reduced affinity energy as well as RMSD less than 2.5. In the same way, a new way of visualizing the Bindscan results using a heatmap chart.
Finally, using the new metric and the new way of visualizing the results, it has been repeated the search for mutants with the fixed W394F mutation and given that the results obtained were not entirely satisfactory, the search for mutants has begun, starting with also of the wild type.



Biarnés Fontal, Xevi


IQS SE - Undergraduate Program in Biotechnology