Degradation of glial fibrillary acidic protein by a calcium dependent proteinase: an electroblot study.

In situ and in vitro degradation of glial fibrillary acidic (GFA) protein in mouse spinal cord was examined with electroblots stained for GFA protein by the peroxidase anti-peroxidase method. Non-degraded, intact GFA protein had a molecular weight of 48 Kdaltons and isoelectric points ranging from pH 5.8 to 6.4. The molecular weights of immunoreactive degradation products ranged from 47 to 28 Kdaltons. All of the degradation products had acid shifted isoelectric points (pH 5.8-5.2). Degradation was prevented by chelating calcium with EGTA. In contrast to in situ degradation, degradation in vitro with 3 mM CaCl2 occurred at a faster rate. The effect of pH and temperature on the degradation process were determined by incubating homogenized spinal cords in 3 mM CaCl2 solutions varying in pH from 4 to 10 and at 4, 37, and 60 degrees C. The greatest number of immunoreactive bands with the lowest molecular weights occurred at pH 8 and 37 degrees C. The results suggest that turnover of glial filaments is in part controlled by a calcium dependent proteinase active near neutral pH similar to that postulated for neurofilament turnover.