The subunit structure and stability of conglutin δ, a sulphurrich protein from the seeds of Lupinus angustifolius L.

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Abstract
The oligomeric structure and stability of conglutin δ, the 2S sulphur‐rich lupin seed protein, has been studied. Molecular weights were determined by sedimentation equilibrium methods in the analytical ultracentrifuge. Conglutin δ2 (Mr 14 000, 2S), the most abundant form, was composed of one large (˜9600) and one small polypeptide chain linked by disulphide bonds. The minor oligomeric form, conglutin δ1 (Mr 28 000, 2.8S), was a disulphide‐linked dimer of conglutin δ2. Each form was capable of reversible association and at low ionic strength (neutral pH), the conglutin δ1 momomer associated to a homogeneous dimer (Mr 56 000, 4.1S). Calculated frictional ratios (f/f0 ˜ 1.28–1.49) suggested that the three different forms were asymmetric. Spectral studies (ORD/CD) showed that conglutins δ1 and δ2 were rich in alpha‐helix (˜38%) unlike the major 7S and 11S legume seed storage proteins. The helical structure was unusually stable both to heat and chemical denaturants; below 60°C (at neutral pH) the helix remained unaffected and only partial denaturation occurred at higher temperatures. Nevertheless, complete denaturation was achieved (at 20°C) in high concentrations of guanidine hydrochloride (greater than 6.5 M). The stability was due to the presence of disulphide cross‐links; with the addition of reducing agent, as little as 1 M guanidine hydrochloride (GuHCl) was sufficient for denaturation of the helical structure. Titration experiments showed a single buried tyrosine (pK 11.4) which could be exposed (pK 10.2) in 6 M GuHCl.