Electrophoretic Studies of Lysine-extracted Plasminogen

hhhough both Tiselius moving boundary electrophoretic and zone electrophoretic techniques have been employed to evaluate the purity of plasminogen (l-11), the interpretation of results can be difficult if protein-protein interactions are involved (12).Recently Kline and Fishman (6) were able to obtain human plasminogen of exceptionally high activity by combining the Kline acid extraction technique (13) with the lysine extraction procedure.The authors indicated that the purified product was homogeneous' according to both moving boundary electrophoretic and ultracentrifugal patterns.They found, however, that the electrophoretic patterns did not change with wide variations in specific activities of the products; and they concluded that the analyses were not decisive as a criterion of plasminogen purity.We have noticed in other published electrophoretic patterns of purified plasminogen (1, 4, 5, 8, 9) as well as in our present investigation a lack of enantiography between boundaries from the ascending and descending limbs of the cell; sometimes skewing or other anomalies were observed.This suggests that "purified plasminogen" may involve proteinprotein interaction.The lysine extraction method has been used in the isolation of plasminogen by several workers (6, 7, 14, 15).While the method of Slotta, Michl, and Santos (7) and its various modifications are milder and the products are more soluble at high pH, the Kline-Fishman preparation appeared to possess the highest biological activity.In this report plasminogen samples prepared by the lysine extraction methods have been used to demonstrate the abnormal behavior in free electrophoresis.This information illuminates, at least in part, the cause of complicat.ioninvolved in the purification and characterization of the enzyme precursor.EXPERIMENTAL PROCEDURE Preparation of Lys-ine-extracted Samples-The samples used were prepared as described below.Each sample has been assigned a number for convenience of discussion.Sample 1 was prepared according to Kline and Fishman's method (6) from Cohn's Fraction III of human plasma.2Sample 2 was prepared from fresh bovine plasma by the Jackson-Mertz method ( 16).The crude plasminogen thus obtained was subjected to further purification by Kline and Fishman's met,hod (6).* Supported by a grant from G. D. Searle and Company. 1 Since no electrophoretic patterns or detailed description of experimental conditions were given in the paper, the meaning of the term "homogeneous" is not clear.2 The human plasma fraction (Cohn's Fraction III) was obtained from E. R. Squibb and Sons through the courtesy of