Expression of human parathyroid hormone in Escherichia coli.
Human parathyroid hormone (hPTH) is a peptide hormone consisting of 84 amino acids. Using the expression plasmid pKK223-3 with the strong tacpromoter, we have produced a variant of hPTH in E. coli. From the expression plasmid construct the expected product was hPTH with an N-terminal extension of Met-Gly. The peptide was extracted from E. coli cells and purified by high performance liquid chromatography. In two different gel electrophoresis systems including identification by immunoblotting the product behaved exactly as an hPTH standard. N-terminal amino acid sequence analysis of the purified product showed traces of Gly-hPTH. At least 90% of the expressed product was N-terminally blocked, suggesting the presence of N-formyl-methionine. This variant of hPTH did not stimulate adenylate cyclase activity in rat osteosarcoma cell membranes.
Høgset
A
A
Institue of Medical Biochemistry, University of Oslo, Norway.
Blingsmo
O R
OR
Gautvik
V T
VT
Saether
O
O
Jacobsen
P B
PB
Gordeladze
J O
JO
Alestrøm
P
P
Gautvik
K M
KM
D000262:Adenylyl Cyclases / Q000378:metabolism; D000595:Amino Acid Sequence; D003001:Cloning, Molecular / Q000379:methods*; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics*; D015870:Gene Expression; D005796:Genes; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010281:Parathyroid Hormone / Q000235:genetics* / Q000302:isolation & purification / Q000494:pharmacology; D010957:Plasmids; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000494:pharmacology; D015183:Restriction Mapping
D010281:Parathyroid Hormone; D011994:Recombinant Proteins; D004247:DNA; D000262:Adenylyl Cyclases