Biological, immunological and biochemical characterization of cleaved prolactin generated by lactating mammary gland.
We have previously shown that rat prolactin is proteolytically cleaved in its loop by peripheral tissues of the rat. Of the tissues examined to date, lactating mammary gland exhibits the highest prolactin-cleaving activity. The objective of this study was to characterize cleaved prolactin, biologically, immunologically and chemically. By modifying an established analytical method, we were able to generate large (micrograms) amounts of cleaved rat prolactin from cell fractions of rat mammary gland which could then be assayed for biological and immunological activity relative to intact hormone. The cleaved product showed no significant difference relative to the intact rat prolactin when assayed for its ability to compete with 125I-labelled ovine prolactin for the prolactin receptor and for its ability to stimulate the proliferation of rat Nb2 lymphoma cells. Cleaved rat prolactin, however, did show a 50-60% reduction in activity relative to intact rat prolactin when assayed by radioimmunoassay. Using Edman degradation and partial amino acid analysis, we determined that the second N-terminus of the cleaved rat prolactin begins at amino acid 149. The divergence of biological and immunological activity produced by proteolytic cleavage in the loop of rat prolactin suggests that biological and immunological sites differ in location. The possible physiological implications of a cleaved rat prolactin molecule generated by target tissue with maintained biological activity and reduced immunological activity are discussed.
Vick
R S
RS
Department of Physiology, Medical College of Virginia, Virginia Commonwealth University, Richmond 23298.
Wong
V L
VL
Witorsch
R J
RJ
D000595:Amino Acid Sequence; D000818:Animals; D005260:Female; D006868:Hydrolysis; D007118:Immunoassay; D007700:Kinetics; D007774:Lactation / Q000378:metabolism*; D008321:Mammary Glands, Animal / Q000378:metabolism*; D008969:Molecular Sequence Data; D011247:Pregnancy; D011388:Prolactin / Q000302:isolation & purification / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D011981:Receptors, Prolactin / Q000378:metabolism
D011981:Receptors, Prolactin; D011388:Prolactin