THERMODYNAMICS OF INTERACTION BETWEEN RECOMBINANT HUMAN LYSOSOMAL alfa-GLUCOSIDASE AND PHARMACOLOGICAL CHAPERONES

Background : Glycogen storage disease type II or Pompe disease is a rare inherited metabolic. Pompe disease is caused by mutations in the gene that encodes the lysosomal hydrolase acid a-glucosidase (GAA). GAA deficiency results in lysosomal glycogen accumulation and cellular dysfunction in multiple tissues, particularly skeletal and cardiac muscle . Enzyme replacement therapy (ERT) with recombinant human GAA (rhGAA; Myozyme, Genzyme Corporation, Cambridge, MA) is the only approved treatment for Pompe disease. Recently, it has been demonstrated that pharmacological chaperones, small molecules that specifically bind and stabilize target proteins, can also enhance the trafficking and cellular activity of a number of different wild-type and mutant forms of lysosomal hydrolases. A general strategy for lysosomal storage disorder is also to search for small pharmacological chaperones that are able to bind the active site of mutant enzymes and to facilitate the folding and transport process. These molecules have the ability to improve the trafficking of the protein between the endoplasmic reticulum (ER) and the lysosome. Various inhibitors derived from deoxynojirimycin (DNJ) have been evaluated as pharmacological chaperones, in different lysosomal storage disorders . Methods : We examined the binding affinity of pharmacological chaperones such as 1-deoxynojirimycin (DNJ) and N-acetylcysteine (NAC) to the rhGAA, Myozyme by Isothermal Titration Calorimetry. We studied thermal stability of the enzyme in the absence and presence of ligands at neutral and acidic pH by Differential Scanning Calorimetry Results : ITC results allowed a complete thermodynamic characterization of the interaction between the studied small ligand and the enzyme while the DSC results gave important information on the enzyme conformational stability as a function of pH.