Solution structure of the α‐subunit of human chorionic gonadotropin

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Abstract
The three‐dimensional solution structure of the α‐subunit in the α,β heterodimeric human chorionic gonadotropin (hCG), deglycosylated with endo‐β‐N‐acetylglucosaminidase‐B (dg‐αhCG), was determined using 2D homonuclear and 2D heteronuclear 1H, 13C NMR spectroscopy at natural abundance in conjunction with the program package x plor. The distance geometry/simulated annealing protocol was modified to allow for the efficient modelling of the cystine knot motif present in αhCG. The protein structure was modelled with 620 interproton distance restraints and the GlcNAc residue linked to Asn78 was modelled with 30 protein‐carbohydrate and 3 intraresidual NOEs. The solution structure of dg‐αhCG is represented by an ensemble of 27 structures. In comparison to the crystal structure of the dimer, the solution structure of free dg‐αhCG exhibits: (a) an increased structural disorder (residues 33–57); (b) a different backbone conformation near Val76 and Glu77; and (c) a larger flexibility. These differences are caused by the absence of the interactions with the β‐subunit. Consequently, in free dg‐αhCG, compared to the intact dimer, the two hairpin loops 20–23 and 70–74 are arranged differently with respect to each other. The β‐GlcNAc(78) is tightly associated with the hydrophobic protein‐core in between the β‐hairpins. This conclusion is based on the NOEs from the axial H1, H3, H5 atoms and the N‐acetyl protons of β‐GlcNAc(78) to the protein‐core. The hydrophobic protein‐core between the β‐hairpins is thereby shielded from the solvent.