Full-length chicken parathyroid hormone. Biosynthesis in Escherichia coli and analysis of biologic activity

J Biol Chem. 1991 Feb 25;266(6):3709-14.

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.

MeSH terms

  • Animals
  • Chickens
  • Chromatography, High Pressure Liquid
  • Cyclic AMP / metabolism
  • Electrophoresis, Polyacrylamide Gel
  • Escherichia coli / genetics*
  • Factor Xa / pharmacology
  • Gene Expression Regulation, Bacterial
  • Genes, Bacterial
  • Immunohistochemistry
  • Parathyroid Hormone / biosynthesis*
  • Parathyroid Hormone / isolation & purification
  • Parathyroid Hormone / metabolism
  • Plasmids
  • Radioligand Assay
  • Rats
  • Tumor Cells, Cultured

Substances

  • Parathyroid Hormone
  • Cyclic AMP
  • Factor Xa