Purification of a candidate gonadotrophin surge attenuating factor from human follicular fluid (Journal article)

Pappa, A./ Seferiadis, K./ Fotsis, T./ Shevchenko, A./ Marselos, M./ Tsolas, O./ Messinis, I. E.

Gonadotrophin surge attenuating factor (GnSAF) is a new non-steroidal ovarian substance, different from inhibin, which attenuates the pre-ovulatory luteinizing hormone (LH) surge in superovulated women. Human follicular fluid (FF) was used as a source for the isolation of GnSAF, the activity of which was monitored in an in-vitro pituitary bioassay. Primary rat pituitary cells were incubated with test substances for 48 h and subsequently washed and incubated with 0.1 micromol/l gonadotrophin releasing hormone (GnRH) plus test substances for 4 h. GnSAF activity was expressed as the reduction of GnRH-induced LH secretion in the 4 h incubation. GnSAF was purified from 250 ml of FF which was heat-treated at 80 degrees C for 5 min. Heparin-sepharose chromatography, Con-A sepharose chromatography, reversed-phase high-performance liquid chromatography (HPLC) and preparative native gel electrophoresis were used for GnSAF fractionation. Using these purification steps, we have obtained an apparently homogeneous preparation that stains as a single band on sodium dodecyl sulphate (SDS)-polyacrylamide gel electrophoresis. GnSAF has an apparent molecular weight of 12.5 kDa and was identified by amino acid sequence (mass spectrometry) to be the C-terminal fragment of human serum albumin.
Institution and School/Department of submitter: Πανεπιστήμιο Ιωαννίνων. Σχολή Επιστημών Υγείας. Τμήμα Ιατρικής
Keywords: Amino Acid Sequence,Animals,Biological Assay,Cells, Cultured,Chromatography, High Pressure Liquid,Electrophoresis, Polyacrylamide Gel,Female,Follicular Fluid/*chemistry,Glycosylation,Gonadal Hormones,Gonadal Steroid Hormones/isolation & purification,Gonadotropin-Releasing Hormone/pharmacology,Hot Temperature,Humans,Luteinizing Hormone/secretion,Molecular Sequence Data,Pituitary Gland/drug effects,Proteins/chemistry/*isolation & purification/pharmacology,Rats,Sequence Homology
URI: http://olympias.lib.uoi.gr/jspui/handle/123456789/18883
ISSN: 0268-1161
Link: http://www.ncbi.nlm.nih.gov/pubmed/10357957
Appears in Collections:Άρθρα σε επιστημονικά περιοδικά ( Ανοικτά)

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