Endocrine Abstracts (2004) 7 P32

Urinary human chorionic gonadotrophin (hCG) associated proteins: isolation and characterisation by peptide identification mass spectrometry

RK Iles1, S Malatos1, SA Butler1, H Neubert2 & AT Kicman2

1Williamson Laboratory for Molecular Oncology, Department of Obstetrics and Gynaecology, St Bartolomew's Hospital, London, UK; 2Drug Control Centre, Department of Forensic Science and Drug Monitoring, King's College London, UK.

Proteins associated with pregnancy urine and urinary-derived hCG preparations have been associated with stimulation of Candida albicans, induction of apoptosis in Kaposi's sarcoma, and inhibition of HIV replication. The active components have been previously reported to be 18 kDa, 15-30 kDa and 2-4 kDa proteins, and suggested in one study to be a ribonuclease (1).

The urinary hCG preparation CG-10 (supplied by Sigma Chemical Company Ltd) is known to be particularly rich in these hCG-associated peptides. The preparation was subjected to size exclusion chromatography and analyses of the resultant eluent fractions indicated the presence of unknown material close in elution time to the urinary degradation product of hCG (hCG-beta core fragment). Further analysis of this material, by polyacrylamide gel electrophoresis (PAGE) and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS), showed it to be a complex of at least two proteins. Isolated bands of protein resolved on PAGE were extracted and subjected to tryptic digestion followed by MALDI-TOF MS and electrospray ionisation mass spectrometry. Tryptic mass mapping and Edman N-terminal sequencing revealed the two proteins to be fragments of inter alpha-1 trypsin inhibitor and transforming growth factor beta. Higher mass protein bands on PAGE were complex aggregates of these two fragments.

Now that we have identified these proteins, their possible role in the biological actions ascribed to components associated with hCG and hCG related metabolite preparations can be thoroughly investigated.

1. Butler, S.A. & Iles, R.K. (2003) Ectopic Human Chorionic Gonadotropin beta secretion by epithelial tumors and Human Chorionic Gonadotropin beta-induced apoptosis in Kaposi's sarcoma: Is there a connection? Clin. Cancer Res. 9: 4666-4673

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