The C-1 subunit of alpha-crustacyanin: the de novo phasing of the crystal structure of a 40 kDa homodimeric protein using the anomalous scattering from S atoms combined with direct methods

The previously unknown crystal structure of the C-1 subunit of the caroteno id-binding protein alpha -crustacyanin has been determined using the anomal ous scattering available at 1.77 Angstrom wavelength to determine the parti al structure of the S atoms intrinsic to the native protein. The resulting 'heavy-atom' phases, in conjunction with near-atomic resolution (d(min) = 1 .15 Angstrom) data, were then used to initiate successful structure determi nation using a direct-methods approach. This is, to the authors' knowledge, the first time such a small anomalous signal (similar to 1%) has been used to aid the determination of a macromolecular structure. As well as the str ucture itself, the methods used during data collection and those used in th e elucidation of the sulfur 'heavy-atom' partial structure are described he re. As predicted, the C-1 subunit adopts a tertiary structure typical of th e lipocalin superfamily: an eight-stranded antiparallel beta -barrel with a repeated +1 topology. The beta -barrel has a calyx shape with the two mole cules in the asymmetric unit interacting in such a way that the open ends o f each calyx face each other, although they do not form a single elongated pocket. A comparison of this structure with those of other members of the l ipocalin superfamily has allowed speculation as to the nature of carotenoid binding by the protein.