Example 1
Introduction
Lafora disease (LD) is an autosomal recessive, fatal progressive myoclonus epilepsy caused by the abnormal buildup of insoluble glycogen, called Lafora bodies. Mutations in the gene encoding the protein laforin lead to LD. Laforin is a dual-specificity phosphatase with a carbohydrate-binding module. This enzyme is necessary for proper en metabolism, but its role in the development of LD glycog not yet fully understood.
Objective
In this study, we established a purification scheme to purify recombinant laforin and analyzed laforin to determine whether the monomer or dimer species is more physiologically relevant our ultimate goal is to crystallize laforin to determine its three-dimensional structure and use these insights to understand the enzyme. Human laforin is difficult to purify due to its tendency to be sequestered into inclusion bodies when expressed in E. coli.
Method
Therefore, we cloned the gene for laforin from the Gallus gallus (red rooster) genome into a bacterial expression vector and purified forin from E. coli using a two-step purification procedure. subjected monomeric Gallus gallus laforin to gel electrophoresis, mass spectrometry, dynamic light scattering phosphatase and mainly as a monomer, remains monomeric , and has phosphatase and carbohydrate-binding activity comparable to human laforin.
result & Conclution
Therefore, Gallus gallus laforin is an appropriate to model for human laforin, any insights we gain from it can be applied human laforin. With this information we can move forward in understanding the role of laforin in the body and eventually develop treatment options for LD