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.
Klotz
A V
AV
Glazer
A N
AN
Bishop
J E
JE
Nagy
J O
JO
Rapoport
H
H
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