Structural modelling of human chorion ic gonadotropin: the effect of nicking and glycosylation on the molecular folding and likely epitope recognition by HCG antibodies.

Aim: Human chorionic gonadotropin is one of four members of the glycoprotein hormone family, so named because between 20-30% of the molecular mass is accounted for by glycosylation. In 1994 the crystal structure of hCG was elucidated and since then the model generated has become the accepted structure of hCG and its molecular variants. With the variation in the extent of glycosylation and nicking effectively giving rise to charge fluctuations across the entire molecule we propose that hCG undergoes topological shifts. Thus, the original model does not fully represent the molecule in its native state due to an additional carboxyl terminal peptide (CTP) and glycosylation which is tremendously variable. In addition, nicking of the Keutman loop is also likely to have a pronounced effects on molecular folding. We propose that these shifts may give rise to epitiope changes effecting antibody recognition.
Methods: Using bioinformatic tools we have examined the effect of variable CTP, glycosylation and nicking and we have characterized a panel of variant hCG molecules. Results: We have built a random coil model of the CTP using sequence editor in hyperchem which when attached to the main peptide shows a limited number of thermodynamically stable conformations and provide evidence for the most likely formation. We have also demonstrated the effect of the normal and hyper-glycosylated O-linked sugars on the CTP and reveal not only three dimensional shifts but also potential secondary structural modifications. N-Linked oligosaccaraides (GGG GGM GGGF) and nicking at 4 sites in the Keutman loop also bring about conformational shifts.
Conclusion: We highlight significant changes to the folding in hCG as a result of introducing these molecular changes and suggest the effect that these alterations may have on well characterised antibodies commonly used to recognise hCG.