A Nicked β-Subunit of Human Chorionic Gonadotropin Purified from Pregnancy Urine1

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Abstract
Human chorionic gonadotropin (hCG) is a glycoprotein hormone composed of two dis-simialr subunits (α and β) and normally excreted in urine of pregnant women. An uncommon β-subunit of hCG was purified from fresh early normal pregnancy urine by Sepralyte C8 resin adsorption, Sephadex G-100 column chromatography, and reverse-phase HPLC. SDS-PAGE under non-reducing conditions showed that the apparent molecular weight (39,000) of this β-subunit was extremely similar to that of the native β-subunit, which is known to consist of 145 amino acid residues and carbohydrates. However, SDS-PAGE, under reducing conditions, resulted in two bands with apparent molecular weights of 22,000 and 18,000, indicating that it consisted of two peptide fragments connected with disulfide bridge(s). These two peptide fragments, separated and purified from the reduced and carboxymethylated protein, were subjected to amino acid and N-terminal sequence analyses. It was found that this β-subunit consisted of two polypeptide chains composed of residues 1-47 disulfide-bridged to residues 48-145 of the β-subunit, which may be produced by nicking of the β-subunit at the one site (Gly47-Val48). This β-subunit was termed a nicked β-subunit of hCG (N-hCGβ). It was also found that N-hCG;9 was present in urine as an a(αβ dimer, indicating that an intrachain nicking of this site in the y9-subunit does not inhibit αβ dimer formation. However, this nick in the subunit did reduce a hormonal activity (the stimulatory effect on testosterone production in rat Leydig cells) of hCG, since the activity of hCG reconstituted with N-hCG7β and intact α-subunit was significantly lower than that of intact hCG.

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