Expression of human parathyroid hormone-(1-84) in Escherichia coli as a factor X-cleavable fusion protein

J Biol Chem. 1990 Sep 15;265(26):15854-9.

Abstract

Recombinant human parathyroid hormone (hPTH)-(1-84) was obtained from Escherichia coli using a cleavable fusion protein strategy. The fusion protein contains residues 1-138 of human growth hormone as the amino-terminal region and residues 1-84 of hPTH as the carboxyl-terminal region. A 7-residue linker containing the recognition/cleavage sequence of the site-specific blood coagulation protease activated factor X (factor Xa) joins the two regions. Intact hPTH-(1-84) is released from this fusion protein by cleavage in vitro with factor Xa. The fusion protein was produced at a high level and formed inclusion bodies which allowed it to be easily purified by low speed centrifugation, with a yield of approximately 50 mg/liter of culture. After factor Xa cleavage and high performance liquid chromatography purification, highly purified hPTH was obtained, with a final yield of 1.5-3 mg/liter. Physical and biological characterization of the purified hormone demonstrated that it was intact and active hPTH-(1-84).

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Base Sequence
  • Cell Line
  • Chromatography, High Pressure Liquid
  • Cyclic AMP / metabolism
  • Electrophoresis, Polyacrylamide Gel
  • Escherichia coli / genetics*
  • Factor X / metabolism*
  • Gene Expression
  • Humans
  • Molecular Sequence Data
  • Molecular Weight
  • Osteosarcoma
  • Parathyroid Hormone / genetics*
  • Parathyroid Hormone / isolation & purification
  • Parathyroid Hormone / pharmacology
  • Plasmids
  • Rats
  • Recombinant Fusion Proteins / isolation & purification
  • Recombinant Fusion Proteins / pharmacology
  • Restriction Mapping

Substances

  • Parathyroid Hormone
  • Recombinant Fusion Proteins
  • Factor X
  • Cyclic AMP