Determination of the Glycoforms of Human Chorionic Gonadotropin β-Core Fragment by Matrix-assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry

[摘要] Background: Metabolism of human chorionic gonadotropin (hCG) in the serum and kidney yields the terminal urinary product hCG β-core fragment (hCGβcf), comprising two disulfide-linked peptides (β6-β40 and β55-β92) of which one (β6-β40) retains truncated N-linked sugars. Hyperglycosylated hCGβcf may indicate choriocarcinoma or Down syndrome, but the glycosylation profile of hCGβcf has not been thoroughly evaluated.Methods: hCGβcf, purified from pregnancy urine, was reduced by “on-target” dithiothreitol (DTT) reduction and analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The mass ([M+H]+) of the primary sequence of the glycosylated peptide β6-β40 was subtracted from the m/z values of the discrete peaks observed to give the masses of the carbohydrate moieties. Carbohydrate structure was predicted by sequentially subtracting the masses of the monosaccharide residues corresponding to N-linked carbohydrates of the hCG β-subunit reported in the literature.Results: Mass spectra of hCGβcf revealed a broad triple peak at m/z 8700–11300. After reduction, the triple peak was replaced by a discrete set of peaks between m/z 4156 and 6354. A peak at m/z 4156.8 corresponded to the nonglycosylated peptide (β55-β92). The remaining nine peaks indicated that urinary hCGβcf comprises a set of glycoforms smaller and larger than the trimannosyl core.Conclusions: hCGβcf comprises a wider set of glycoforms than reported previously. Peaks of highest mass indicate evidence of hyperglycosylated carbohydrate moieties. The data support previous reports that hCGβcf oligosaccharides lack sialic acid and galactose residues. No indication was found of a β6-β40 peptide that was entirely devoid of carbohydrate.

[学科分类] 过敏症与临床免疫学