Biosynthesis of human endothelins in transformants expressing cDNAs for human prepro-endothelins.
To understand the biosynthesis of the human ET family, three kinds of cloned cDNA for human prepro-endothelin-1 (prepro-ET-1), prepro-ET-2, and prepro-ET-3 were stably expressed in Chinese hamster ovary cells (CHO-K1). Immunoreactive (ir-) ET polypeptides in the culture media of the transformants were purified by reverse-phase high-performance liquid chromatography (HPLC) coupled with sandwich-type enzyme immunoassay (EIA). Amino acid sequencing and FAB mass spectrometry of the purified ir-ET polypeptides revealed the presence of human big ET-1 (1-38), big ET-2 (1-38), and big ET-3 (1-41)NH2 as intermediate forms. These results directly revealed the biosynthetic pathways of three human ETs at the peptide level.
Kosaka
T
T
Discovery Research Laboratories, Takeda Chemical Industries, Ltd., Ibaraki.
Suzuki
N
N
Ishibashi
Y
Y
Matsumoto
H
H
Itoh
Y
Y
Ohkubo
S
S
Ogi
K
K
Kitada
C
C
Onda
H
H
Fujino
M
M
D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D006224:Cricetinae; D003470:Culture Media; D018076:DNA, Complementary / Q000235:genetics*; D019332:Endothelin-1; D016232:Endothelins / Q000096:biosynthesis* / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D015870:Gene Expression; D006801:Humans; D007124:Immunoenzyme Techniques; D008969:Molecular Sequence Data; D011498:Protein Precursors / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D016339:Spectrometry, Mass, Fast Atom Bombardment; D014162:Transfection; D014170:Transformation, Genetic
D003470:Culture Media; D018076:DNA, Complementary; D019332:Endothelin-1; D016232:Endothelins; D011498:Protein Precursors