Expression and characterization of a recombinant human parathyroid hormone secreted by Escherichia coli employing the staphylococcal protein A promoter and signal sequence.
Human parathyroid hormone (hPTH) is a peptide hormone consisting of 84 amino acids (hPTH(1-84)). Employing the promoter and signal sequence of Staphylococcus aureus-protein A we have expressed hPTH in Escherichia coli. The expressed proteins are excreted to the growth medium, allowing for rapid and easy purification of the desired products. By amino acid sequence analysis and mass spectrometry, we have shown that the major excreted product is correctly processed human identical hPTH(1-84). The purified recombinant hPTH(1-84) stimulates adenylate cyclase activity in rat osteosarcoma cell membranes to exactly the same extent as synthetic parathyroid hormone standards, indicating that the recombinant product has full biological activity.
Høgset
A
A
Institute of Medical Biochemistry, University of Oslo, Blindern, Norway.
Blingsmo
O R
OR
Saether
O
O
Gautvik
V T
VT
Holmgren
E
E
Hartmanis
M
M
Josephson
S
S
Gabrielsen
O S
OS
Gordeladze
J O
JO
Alestrøm
P
P
D000262:Adenylyl Cyclases / Q000378:metabolism; D000595:Amino Acid Sequence; D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics* / Q000254:growth & development; D006801:Humans; D015151:Immunoblotting; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010281:Parathyroid Hormone / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D010957:Plasmids; D011401:Promoter Regions, Genetic; D021382:Protein Sorting Signals / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000494:pharmacology; D015183:Restriction Mapping; D013205:Staphylococcal Protein A / Q000235:genetics*
D010281:Parathyroid Hormone; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; D013205:Staphylococcal Protein A; D000262:Adenylyl Cyclases