Functional study of a recombinant form of human LHbeta-subunit variant carrying the Gly(102)Ser mutation found in Asian populations.

Abstract

Genetic variants of human LH caused by amino acid replacements in the beta-subunit have been demonstrated to affect reproductive function. Occurrence of a G(1502)A substitution in the LHbeta gene leading to Gly(102)Ser replacement of the LHbeta protein has been found to be associated with infertility in the Singapore Chinese population. In the present study, a search for this LHbeta allele from 383 DNA samples from different continents, using a PCR-based strategy, demonstrated its total absence in these populations. Functional properties of the variant (V) (Gly(102)Ser substitution) LHbeta subunit were assessed using a recombinant (r) form of V-LH produced in HEK293 cells, in comparison with wild-type (WT) LH or hCG. The synthesized V-LH was purified by a single step of immunoaffinity chromatography, and it had a molecular weight of 30 kDa as determined by SDS-PAGE. The affinities of the WT-hCG and rV-LH in mouse Leydig tumour (mLT-1) cell LH receptor binding were similar, with K(d) values of 0.140 +/- 0.03 and 0.156 +/- 0.01 nmol/l respectively. Likewise, the effects of WT- and V-rLH preparations on mLT-1 cell cAMP and progesterone production were concentration-dependent and with similar biopotencies. In addition, HEK293 cells expressing the human LH receptor documented similar dose-dependent increases in inositol phosphate production by the two rLH forms. In conclusion, these findings demonstrate that Gly(102)Ser mutation of the LHbeta gene does not affect receptor binding and bioactivity of the hormone, when tested in vitro.