Characterization of Sheep Prolactin.

Prolactin has been obtained from sheep pituitary glands by several methods (1, 2). Cole and Li (3) described a simplified procedure for the preparation of sheep prolactin and its characterization by physicochemical methods (4, 5). The same investigators reported later that prolactin obtained by this method was electrophoretically heterogeneous, but that the three components had essentially equal lactogenic activity (6). Pierce and Carsten (7) observed molecular heterogeneity by submitting prolactin to zone electrophoresis on starch gel, and they showed that only one component passed through a stretched membrane calibrated to be permeable to proteins with molecular weights up to 30,000. Cole (8) has described a method for the further purification of prolactin by ion exchange chromatography on Amberlite IRC-50 columns in urea-containing buffers. Each of the three components thus resolved had equal lactogenic activity and also contained luteotropic activity. Ferguson and Wallace (9), however, reported that the individual components of sheep prolactin separated by starch gel electrophoresis possess markedly different lactogenic activities. Recently, Squire, Starman, and Li (10) described ultracentrifugation experiments which indicate that prolactin prepared by the procedures of Cole and Li (3) contains appreciable amounts of polymeric material which is not in equilibrium with the monomer. Gel filtration on Sephadex G-75 separated a monomer fraction essentially pure by sedimentation characteristics, but exhibiting electrophoretic heterogeneity; i.e. showing five bands by starch gel electrophoresis. In a preliminary communication (11)) we outlined a procedure utilizing chromatography on diethylaminoethyl cellulose for the isolation in substantially pure form of a major component (Component I) of sheep prolactin with markedly greater lactogenic activity than that of accompanying components as determined by the quantitative systemic pigeon crop sac assay. More recently one of our partially purified prolactin preparations, which by electrophoretic criteria contained 85 to 90% of Component I, was found to act as an anabolic agent in man (12), and to mimic the effects of human growth hormone without producing any serious antigenic effects even after repeated dosage over an extended period of time (13). The purpose of this paper is t.o describe in further detail the isolation procedure of Component I and some of its physical and chemical characteristics.