EFFECTS OF DRYING METHODS AND ADDITIVES ON STRUCTURE AND FUNCTION OF ACTIN - MECHANISMS OF DEHYDRATION-INDUCED DAMAGE AND ITS INHIBITION

Limited stability impedes the development of industrial and pharmaceut ical proteins. Dried formulations are theoretically more stable, but t he drying process itself causes structural damage leading to loss of a ctivity after rehydration. Lyophilization is the most common method us ed to dry proteins, but involves freezing and dehydration, which are b oth damaging to protein. We compared an air-drying method to freeze-dr ying to test the hypothesis that terminal dehydration is the critical stress leading to loss of activity. The secondary structure of air-dri ed and freeze-dried actin was analyzed by infrared spectroscopy and re lated to the level of activity recovered from the rehydrated samples. Actin dried by either method in the absence of stabilizers was highly unfolded and the capacity to polymerize was lost upon rehydration, The degree of unfolding was reduced by air-drying or freeze-drying actin with sucrose, and the level of activity recovered upon rehydration inc reased. The addition of dextran to sucrose improved the recovery of ac tivity from freeze-dried, but not air-dried samples. Dextran alone fai led to protect the structure and function of actin dried by either met hod, indicating that proteins are not protected from dehydration-induc ed damage by formation of a glassy matrix. In some cases, recovered ac tivity did not correlate directly with the level of structural protect ion conferred by a particular additive. This result suggests that seco ndary structural protection during drying is a necessary but not suffi cient condition for the recovery of activity from a dried protein afte r rehydration. (C) 1998 Academic Press.