The specific aim of this proposal is to purify to homogeneity acidic Alpha-D-mannosidase from human placenta and bovine kidney. It is anticipated that by employing a combination of chromatographic techniques such as affinity, high-performance anion exchange and/or high performance gel filtration matrices, homogeneous acidic Alpha-D-mannosidase will be purified in large-scale. This will permit the generation of polyclonal and monoclonal antibodies during Phase II. Furthermore, the purified enzyme and suitable antiserum will provide a means for cloning the gene for acidic Alpha-D-mannosidase. The absence this enzyme in the lysosomal storage disorder, mannosidosis, which occurs in both man and cattle, makes it an ideal candidate for studying the feasibility of gene replacement therapy in the animal, prior to attempting it in human subjects.
