EXPRESSION STUDIES OF CLN3 PROTEIN

Expression of the gene for Batten disease (CLN3) was studied in Escher ichia coli and in a cell-free rabbit reticulocyte expression systems, A full-length recombinant fusion CLN3 protein was not produced in the bacterial systems used. However, both N-terminal fragment encompassing 246 amino acids and short C-terminal fragment containing 428-438 amin o acids of the CLN3 protein were successfully overexpressed in bacteri a. Further studies showed that the C-terminal sequence of the CLN3 pro tein corresponding to the 356-438 amino acid residues was responsible for inhibition of protein synthesis in bacteria. The full-length CLN3 gene product was readily synthesized in vitro in the cell-free rabbit reticulocyte expression system. The product obtained, corresponding to core CLN3 protein, showed an approximate molecular weight of 43 kDa. Immunoprecipitation of this product with pAb to 4-19 amino acids of th e CLN3 protein allows us to suggest that CLN3 protein translation star ts at Met-1.