title: Full-length chicken parathyroid hormone. Biosynthesis in Escherichia coli and analysis of biologic activity.
abstract: Chicken parathyroid hormone (cPTH) has been reported to stimulate adrenal steroidogenesis and to have unusual potency on traditional PTH target tissues. To evaluate these properties, chicken PTH-(1-88) has been expressed in Escherichia coli using a plasmid encoding a fusion protein which links together growth hormone, a factor Xa recognition site, and chicken PTH-(1-88). The growth hormone-cPTH fusion protein required the presence of 0.02% sodium dodecyl sulfate to remain in solution and be cleaved by factor Xa. The high performance liquid chromatography-purified recombinant cPTH-(1-88) and chemically synthesized cPTH-(1-34) had similar potency in rat osteosarcoma (ROS 17/2.8) cells, opossum kidney (OK) cells, and dispersed primary chicken kidney cells. The biologic potencies of cPTH-(1-34) and cPTH-(1-88) in radioreceptor binding and cAMP generation in both bone- and kidney-derived cell lines were less than those of human (h)PTH-(1-34). In dispersed chicken kidney cells, cAMP production by cPTH-(1-34) and cPTH-(1-88) was similar to that stimulated by human PTH-(1-34). No stimulation of steroidogenesis could be detected when recombinant chicken PTH-(1-88) was added to dispersed chicken adrenal cells. The biologic activity of recombinant chicken PTH-(1-88) purified from E. coli was comparable with that of chicken PTH-(1-88) expressed by mammalian COS cells. Thus, the full-length chicken PTH did not exhibit enhanced potency, when compared with human PTH in ROS 17/2.8, OK cell lines, and dispersed chicken kidney cells and did not demonstrate the novel steroidogenic action previously reported in adrenal cells. The successful production of chicken PTH-(1-88) will enhance our understanding of the structure-activity relationships for PTH, particularly the sequence-dependent metabolism of the hormone.
authors.affiliation: Endocrine Unit, Massachusetts General Hospital, Boston.
authors.forename: S K
authors.initials: SK
authors.lastname: Lim
authors.forename: T
authors.initials: T
authors.lastname: Gardella
authors.forename: A
authors.initials: A
authors.lastname: Thompson
authors.forename: J
authors.initials: J
authors.lastname: Rosenberg
authors.forename: H
authors.initials: H
authors.lastname: Keutmann
authors.forename: J
authors.initials: J
authors.lastname: Potts
authors.forename: H
authors.initials: H
authors.lastname: Kronenberg
authors.forename: S
authors.initials: S
authors.lastname: Nussbaum
mesh_terms: D000818:Animals; D002645:Chickens; D002851:Chromatography, High Pressure Liquid; D000242:Cyclic AMP / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics*; D015951:Factor Xa / Q000494:pharmacology; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D007150:Immunohistochemistry; D010281:Parathyroid Hormone / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D010957:Plasmids; D011869:Radioligand Assay; D051381:Rats; D014407:Tumor Cells, Cultured
chemical_list: D010281:Parathyroid Hormone; D000242:Cyclic AMP; D015951:Factor Xa