The structure and refinement of apocrustacyanin C₂to 1.3 Å resolution and the search for differences between this protein and the homologous apoproteins A₁and C₁

Abstract

The blue carotenoprotein alpha-crustacyanin of Homarus gammarus lobster carapace is comprised chemically of five 20 kDa subunits. Only two genes for the proteins have been isolated (J. B. C. Findlay, personal communication) and the five apoproteins fall into two sets of homologous proteins based on their chemical properties (CRTC, consisting of apoproteins C(1), C(2) and A(1), and CRTA, consisting of apoproteins A(2) and A(3)). The diffraction quality of apo C(2) has been improved from 2.2 to 1.3 A and its structure solved. The structure is compared with the A(1) and C(1) proteins determined at 1.4 A [Cianci et al. (2001), Acta Cryst. D57, 1219-1229] and 1.15 A, respectively [Gordon et al. (2001), Acta Cryst. D57, 1230-1237] and found to be very similar. Normalized B-factor difference plots per residue of different types were used to try to find chemically modified residues; none were found at these resolutions. It remains possible that the differences between the CRTC proteins result from differences in amidation. By comparison of a crystal grown with glycerol (studied at 1.6 A) and one grown without glycerol (studied at 1.3 A) it was seen that glycerol bound at the astaxanthin site.

Structural characterization of recombinant crustacyanin subunits from the lobsterHomarus americanus

“…2a-2d), with a -barrel made up of two distinct -sheets. Homodimerization is achieved via a close contact between the two -strands of each subunit, similarly to the A 1 (Cianci et al, 2001), C 1 (Gordon et al, 2001) and C 2 (Habash et al, 2004) crystal structures (see Fig. 4 of Gordon et al, 2001).…”

“…The H 1 protein has 181 residues with the typical lipocalin folding of crustacyanins A 1 (Cianci et al, 2001(Cianci et al, , 2002, C 1 (Gordon et al, 2001) and C 2 (Habash et al, 2004) (Figs. 2a-2d), with a -barrel made up of two distinct -sheets.…”

Detailed spectroscopic study of the role of Br and Sr in coloured parts of the Callinectes sapidus crab claw

“…The EXAFS analysis results are in excellent agreement with the findings of Feiters et al (2005aFeiters et al ( , 2005b for these reference compounds, providing thus evidence for Br atoms being trapped in the phenyl and/or phenol rings located in the amino acid residues of the chromophore protein. More specifically, 12 phenylalanine, 14 tyrosine and 2 tryptophan aromatic amino acid residues were reported in various crustacyanin proteins (1GKA, 1I4U, 1S44) in the protein data bank (Berman et al, 2000;Cianci et al, 2002;Gordon et al, 2001;Habash et al, 2004). Incorporation of Br in native bromoperoxidase enzyme from Ascophyllum nodosum by covalent bonding of Br to phenol rings of tyrosine residues has been verified by EXAFS (Feiters et al, 2005).…”

Rapid expansion of pigmentation genes in penaeid shrimp with absolute preservation of function

“…Five distinct carotenoid-binding proteins (CBPs) were originally identified from the European lobster (Homarus gammarus) and these proteins were grouped into the broad classes CRCN-A and CRCN-C based upon amino acid composition, electrophoretic mobility and peptide mapping (Cheesman et al, 1966;Quarmby et al, 1977). The protein forms a dimer composed of two 20 kDa CRCN subunits and two astaxanthin (Axn) molecules, known as β-crustacyanin, which in turn form a large multimeric complex, called α-crustacyanin, composed of eight dimeric β-crustacyanin subunits Cheesman et al, 1966;Habash et al, 2004;Helliwell, 2010;Zagalsky and Cheesman, 1963).…”

“…The full protein sequence was identified by protein purification and direct sequencing methods for the predominant CRCN A and C subunits (Keen et al, 1991a,b). It is thought that post-translational modifications, for example amidation or glycosylation, are responsible for the differences in electrophoretic mobility of subunits (Habash et al, 2004). In the American lobster (Homarus americanus) only two protein isoforms H1 and H2 have been identified (Zagalsky and Tidmarsh, 1985).…”