Structure of lobster apocrustacyanin A₁ using softer X-rays

The molecular basis of the camouflage colouration of marine crustacea is often provided by carotenoproteins. The blue colour of the lobster carapace, for example, is intricately associated with a multimacromolecular 16-mer complex of protein subunits each with a bound astaxanthin molecule. The protein subunits of crustacyanin fall into two distinct subfamilies, CRTC and CRTA. Here, the crystal structure solution of the A₁ protein of the CRTC subfamily is reported. The problematic nature of the structure solution of the CRTC proteins (both C₁ and A₁) warranted consideration and the development of new approaches. Three putative disulfides per protein subunit were likely to exist based on molecular-­homology modelling against known lipocalin protein structures. With two such subunits per crystallographic asymmetric unit, this direct approach was still difficult as it involved detecting a weak signal from these sulfurs and suggested the use of softer X-rays, combined with high data multiplicity, as reported previously [Chayen et al. (2000), Acta Cryst. D56, 1064–1066]. This paper now describes the structure solution of CRTC in the form of the A₁ dimer based on use of softer X-­rays (2 Å wavelength). The structure solution involved a xenon derivative with an optimized xenon L_I edge signal and a native data set. The hand of the xenon SIROAS phases was determined by using the sulfur anomalous signal from a high-multiplicity native data set also recorded at 2 Å wavelength. For refinement, a high-resolution data set was measured at short wavelength. All four data sets were collected at 100 K. The refined structure to 1.4 Å resolution based on 60 276 reflections has an R factor of 17.7% and an R_free of 22.9% (3137 reflections). The structure is that of a typical lipocalin, being closely related to insecticyanin, to bilin-binding protein and to retinol-binding protein. This A₁ monomer or dimer can now be used as a search motif in the structural studies of the oligomeric forms α- and β-crustacyanins, which contain bound astaxanthin molecules.