title: In vitro attachment of bilins to apophycocyanin. II. Determination of the structures of tryptic bilin peptides derived from the phycocyanobilin adduct.
abstract: In vitro reaction of phycocyanobilin (PCB) with apophycocyanin results in the specific addition of the bilin to two of the cysteinyl residues, alpha-Cys-84 and beta-Cys-82, which normally function in PCB attachment (Arciero, D. M., Bryant, D. A., and Glazer, A. N. (1988) J. Biol. Chem. 263, 18343-18349). These bilin binding sites are designated alpha-1 and beta-1, respectively. Tryptic digestion of the apophycocyanin-PCB adduct releases two major bilin peptides, alpha-1 mesobiliverdin (MBV) and beta-1 MBV, which encompass the two bilin-binding sites. These peptides were examined by 1H NMR and fast atom bombardment mass spectroscopies. The NMR spectra show that the bilin is attached to each peptide through a thioether linkage identical to the linkage observed in the corresponding tryptic peptides, alpha-1 PCB and beta-1 PCB, derived from the natural product, C-phycocyanin. However, the NMR spectra of the adduct peptides lack the resonances corresponding to protons at positions C2 and C3 of ring A seen in the spectra of the alpha-1 PCB and beta-1 PCB peptides. Fast atom bombardment mass spectroscopy shows the masses of the alpha-1 MBV and beta-1 MBV peptides to be 2 atomic mass units lower than those of the alpha-1 PCB and beta-1 PCB peptides, respectively. Comparison of the bilin portion of the NMR spectra of the alpha-1 MBV and beta-1 MBV peptides to the NMR spectra of PCB and mesobiliverdin confirms that the bilin of the two adduct peptides resembles mesobiliverdin in having an extra double bond in the C2-C3 position of ring A. These results show that the major bilin products arising from the reaction of PCB with apophycocyanin differ from the bilins present in C-phycocyanin. The relevance of these results to the biosynthetic pathway for the attachment of tetrapyrroles to phycobiliproteins is discussed.
authors.affiliation: Department of Microbiology and Immunology, University of California, Berkeley 94720.
authors.forename: D M
authors.initials: DM
authors.lastname: Arciero
authors.forename: J L
authors.initials: JL
authors.lastname: Dallas
authors.forename: A N
authors.initials: AN
authors.lastname: Glazer
mesh_terms: D001059:Apoproteins; D001664:Biliverdine / Q000031:analogs & derivatives; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D010446:Peptide Fragments / Q000032:analysis; D052980:Phycobilins; D010798:Phycocyanin / Q000031:analogs & derivatives*; D010860:Pigments, Biological; D011758:Pyrroles; D045725:Tetrapyrroles; D014357:Trypsin
chemical_list: D001059:Apoproteins; D010446:Peptide Fragments; D052980:Phycobilins; D010860:Pigments, Biological; D011758:Pyrroles; D045725:Tetrapyrroles; C057786:apophycocyanin; D010798:Phycocyanin; C011069:phycocyanobilin; C040381:mesobiliverdin; D014357:Trypsin; D001664:Biliverdine