Isoelectric charge of recombinant human follicle-stimulating hormone isoforms determines receptor affinity and in vitro bioactivity.
Recombinant human FSH (rhFSH) was obtained by expressing the human FSH alpha- and beta-subunit complementary DNAs in the chinese hamster ovary cell line. Isoforms of rhFSH were resolved into specific isoelectric (pI) fractions by chromatofocusing. rhFSH isoforms ranged from pI 3.0-5.5 with a modal value of pI 4.2. Analysis of the biological activity of specific pI isoforms of rhFSH was undertaken using both the rat granulosa cell aromatase (in vitro) bioassay and a RRA. More acidic isoforms (e.g. pI 3.5) showed significantly lower affinity (P < 0.05) for rat testicular FSH receptors than did the less acidic isoforms (e.g. pI 4.8). Consistent with the receptor binding affinity data, the more acidic fractions resulted in significantly less activation (P < 0.05) of rat granulosa cell aromatase activity, as measured by estrogen production, than did the less acidic isoforms. The observed bioactivities and their correlation with the pI values of the rhFSH isoforms are consistent with observations of differing bioactivities seen in both pituitary and urinary FSH isoforms. These results demonstrate that rhFSH, made in the chinese hamster ovary cell line, is both biologically active and has isoform profiles, and presumably carbohydrate structures, that closely resemble those seen in natural hFSH.
Cerpa-Poljak
A
A
Garvan Institute of Medical Research, St. Vincents Hospital, Darlinghurst, Sydney, Australia.
Bishop
L A
LA
Hort
Y J
YJ
Chin
C K
CK
DeKroon
R
R
Mahler
S M
SM
Smith
G M
GM
Stuart
M C
MC
Schofield
P R
PR
D000818:Animals; D001141:Aromatase / Q000378:metabolism; D015153:Blotting, Western; D016466:CHO Cells; D006224:Cricetinae; D004355:Drug Stability; D005260:Female; D005640:Follicle Stimulating Hormone / Q000737:chemistry* / Q000235:genetics / Q000494:pharmacology; D015870:Gene Expression; D006107:Granulosa Cells / Q000187:drug effects; D006801:Humans; D007525:Isoelectric Focusing; D007526:Isoelectric Point; D008297:Male; D011869:Radioligand Assay; D051381:Rats; D017208:Rats, Wistar; D011994:Recombinant Proteins / Q000737:chemistry; D014162:Transfection
D011994:Recombinant Proteins; D005640:Follicle Stimulating Hormone; D001141:Aromatase