Phycobiliprotein-bilin linkage diversity. II. Structural studies on A- and D-ring-linked phycoerythrobilins.
The discovery that the phycocyanobilin group attached to Cys-155 of the beta subunit of C-phycocyanin is D-ring linked (Bishop, J. E., Lagarias, J. C., Nagy, J. O., Schoenleber, R. W., Rapoport, H., Klotz, A. V., and Glazer, A. N. (1986) J. Biol. Chem. 261, 6790-6796) prompted examination of the linkage mode for phycoerythrobilin (PEB) groups attached at the corresponding position in other biliproteins. Appropriate small peptides were obtained by exhaustive enzymatic digestion of Porphyridium cruentum R-phycocyanin (peptide R-PC beta-2TP PEB) and B-phycoerythrin (peptide B-PE beta-2TP PEB). These peptides had the following structures R-PC beta-2TP PEB Gly-Asp-Cys(PEB)-Ser-Ser B-PE beta-2TP PEB Cys(PEB)-Thr-Ser. The spectroscopic and chemical properties of these peptides were compared with those of P. cruentum B-phycoerythrin peptide alpha-1 PEB, Cys(PEB)-Tyr-Arg, in which the bilin is A-ring linked (Schoenleber, R. W., Leung, S.-L., Lundell, D. J., Glazer, A. N., and Rapoport, H. (1983) J. Am. Chem. Soc. 105, 4072-4076). The PEB groups in peptides R-PC beta-2TP PEB and B-PE beta-2TP PEB were shown to be D-ring linked on the basis of the following criteria. Secondary ion mass spectrometry showed the bilins in these peptides and in alpha-1 PEB to have the same mass. The 18'-CH3, 18'-H, and 15-H resonances in the 1H NMR spectra of R-PC beta-2TP PEB and B-PE beta-2TP PEB appear significantly upfield from the corresponding thioether-linked ring A resonances seen in the spectrum of peptide alpha-1 PEB. The CD spectra of the two former peptides showed a strong positive Cotton effect at 300 nm. Such a Cotton effect is absent from the CD spectrum of peptide alpha-1 PEB and those of other A-ring-linked PEB peptides. Refluxing in methanol led to a near-quantitative release of PEB from alpha-1 PEB but no release from R-PC beta-2TP PEB and less than 20% release from B-PE beta-2TP PEB. In conjunction with earlier studies, these results show that distinctive amino acid sequences are found about the attachment sites for A-ring-linked, D-ring-linked, and dilinked (A- and D-ring-linked) bilins on the alpha and beta subunits of cyanobacterial and red algal phycobiliproteins and that the mode of linkage can be correctly predicted from inspection of the amino acid sequence.
A V
AV
Klotz
A N
AN
Glazer
J E
JE
Bishop
J O
JO
Nagy
H
H
Rapoport
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D002942:Circular Dichroism; D045342:Light-Harvesting Protein Complexes; D046911:Macromolecular Substances; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D010446:Peptide Fragments / Q000032:analysis; D010799:Phycoerythrin / Q000302:isolation & purification / Q000378:metabolism; D010860:Pigments, Biological / Q000378:metabolism*; D010940:Plant Proteins / Q000302:isolation & purification / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D000461:Rhodophyta / Q000378:metabolism*
D000596:Amino Acids; D045342:Light-Harvesting Protein Complexes; D046911:Macromolecular Substances; D010446:Peptide Fragments; D010860:Pigments, Biological; D010940:Plant Proteins; D010799:Phycoerythrin