Irradiation Inactivation Analysis of Acetylcholinesterase and the Effect of Buffer Salts

Loss of function of membrane-bound proteins after bombardment with ionizing radiation has been used to obtain information about the size of such proteins. Human erythrocyte ghosts are a convenient source of acetylcholinesterase, a membrane-bound enzyme which has been studied with this technique and so could be used as an internal standard for calibration with the high doses required. After freeze-drying from buffers containing Tris, Hepes, or Bicine, irradiation of erythrocyte ghosts with increasing doses of 16-MeV electrons gave size estimates for acetylcholinesterase of between 56,000 and 80,000 molecular weight. These values are similar to those that have been reported in the literature. However, erythrocyte ghosts freezedried from phosphate buffer gave larger size estimates for acetylcholinesterase of about twice that found in the other buffers. This value was close to the molecular weight of 154,000 reported for the purified enzyme. The different sizes did not appear to be due to different ionic stre...