Skip to main content
NCBI home page
Protein Science : A Publication of the Protein Society logoLink to Protein Science : A Publication of the Protein Society
. 1993 Jul;2(7):1114–1125. doi: 10.1002/pro.5560020706

Primary structure of a photoactive yellow protein from the phototrophic bacterium Ectothiorhodospira halophila, with evidence for the mass and the binding site of the chromophore.

J J Van Beeumen 1, B V Devreese 1, S M Van Bun 1, W D Hoff 1, K J Hellingwerf 1, T E Meyer 1, D E McRee 1, M A Cusanovich 1
PMCID: PMC2142427  PMID: 8358295

Abstract

The complete amino acid sequence of the 125-residue photoactive yellow protein (PYP) from Ectothiorhodospira halophila has been determined to be MEHVAFGSEDIENTLAKMDDGQLDGLAFGAIQLDGDGNILQYNAAEGDITGRDPKEVIGKNFFKDVAP+ ++ CTDSPEFYGKFKEGVASGNLNTMFEYTFDYQMTPTKVKVHMKKALSGDSYWVFVKRV. This is the first sequence to be reported for this class of proteins. There is no obvious sequence homology to any other protein, although the crystal structure, known at 2.4 A resolution (McRee, D.E., et al., 1989, Proc. Natl. Acad. Sci. USA 86, 6533-6537), indicates a relationship to the similarly sized fatty acid binding protein (FABP), a representative of a family of eukaryotic proteins that bind hydrophobic molecules. The amino acid sequence exhibits no greater similarity between PYP and FABP than for proteins chosen at random (8%). The photoactive yellow protein contains an unidentified chromophore that is bleached by light but recovers within a second. Here we demonstrate that the chromophore is bound covalently to Cys 69 instead of Lys 111 as deduced from the crystal structure analysis. The partially exposed side chains of Tyr 76, 94, and 118, plus Trp 119 appear to be arranged in a cluster and probably become more exposed due to a conformational change of the protein resulting from light-induced chromophore bleaching. The charged residues are not uniformly distributed on the protein surface but are arranged in positive and negative clusters on opposite sides of the protein. The exact chemical nature of the chromophore remains undetermined, but we here propose a possible structure based on precise mass analysis of a chromophore-binding peptide by electrospray ionization mass spectrometry and on the fact that the chromophore can be cleaved off the apoprotein upon reduction with a thiol reagent. The molecular mass of the chromophore, including an SH group, is 147.6 Da (+/- 0.5 Da); the cysteine residue to which it is bound is at sequence position 69.

Full Text

The Full Text of this article is available as a PDF (2.6 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Alpers D. H., Strauss A. W., Ockner R. K., Bass N. M., Gordon J. I. Cloning of a cDNA encoding rat intestinal fatty acid binding protein. Proc Natl Acad Sci U S A. 1984 Jan;81(2):313–317. doi: 10.1073/pnas.81.2.313. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Cotter R. J. Plasma desorption mass spectrometry: coming of age. Anal Chem. 1988 Jul 1;60(13):781A–793A. doi: 10.1021/ac00164a002. [DOI] [PubMed] [Google Scholar]
  3. Holden H. M., Rypniewski W. R., Law J. H., Rayment I. The molecular structure of insecticyanin from the tobacco hornworm Manduca sexta L. at 2.6 A resolution. EMBO J. 1987 Jun;6(6):1565–1570. doi: 10.1002/j.1460-2075.1987.tb02401.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Hunt C. R., Ro J. H., Dobson D. E., Min H. Y., Spiegelman B. M. Adipocyte P2 gene: developmental expression and homology of 5'-flanking sequences among fat cell-specific genes. Proc Natl Acad Sci U S A. 1986 Jun;83(11):3786–3790. doi: 10.1073/pnas.83.11.3786. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. James G., Olson E. N. Fatty acylated proteins as components of intracellular signaling pathways. Biochemistry. 1990 Mar 20;29(11):2623–2634. doi: 10.1021/bi00463a001. [DOI] [PubMed] [Google Scholar]
  6. Jones T. A., Bergfors T., Sedzik J., Unge T. The three-dimensional structure of P2 myelin protein. EMBO J. 1988 Jun;7(6):1597–1604. doi: 10.1002/j.1460-2075.1988.tb02985.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Li E., Demmer L. A., Sweetser D. A., Ong D. E., Gordon J. I. Rat cellular retinol-binding protein II: use of a cloned cDNA to define its primary structure, tissue-specific expression, and developmental regulation. Proc Natl Acad Sci U S A. 1986 Aug;83(16):5779–5783. doi: 10.1073/pnas.83.16.5779. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Marshall C. J. Protein prenylation: a mediator of protein-protein interactions. Science. 1993 Mar 26;259(5103):1865–1866. doi: 10.1126/science.8456312. [DOI] [PubMed] [Google Scholar]
  9. Meyer T. E., Fitch J. C., Bartsch R. G., Tollin G., Cusanovich M. A. Soluble cytochromes and a photoactive yellow protein isolated from the moderately halophilic purple phototrophic bacterium, Rhodospirillum salexigens. Biochim Biophys Acta. 1990 Apr 26;1016(3):364–370. doi: 10.1016/0005-2728(90)90170-9. [DOI] [PubMed] [Google Scholar]
  10. Meyer T. E. Isolation and characterization of soluble cytochromes, ferredoxins and other chromophoric proteins from the halophilic phototrophic bacterium Ectothiorhodospira halophila. Biochim Biophys Acta. 1985 Jan 23;806(1):175–183. doi: 10.1016/0005-2728(85)90094-5. [DOI] [PubMed] [Google Scholar]
  11. Meyer T. E., Kamen M. D. New perspectives on c-type cytochromes. Adv Protein Chem. 1982;35:105–212. doi: 10.1016/s0065-3233(08)60469-6. [DOI] [PubMed] [Google Scholar]
  12. Meyer T. E., Tollin G., Causgrove T. P., Cheng P., Blankenship R. E. Picosecond decay kinetics and quantum yield of fluorescence of the photoactive yellow protein from the halophilic purple phototrophic bacterium, Ectothiorhodospira halophila. Biophys J. 1991 May;59(5):988–991. doi: 10.1016/S0006-3495(91)82313-X. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Meyer T. E., Tollin G., Hazzard J. H., Cusanovich M. A. Photoactive yellow protein from the purple phototrophic bacterium, Ectothiorhodospira halophila. Quantum yield of photobleaching and effects of temperature, alcohols, glycerol, and sucrose on kinetics of photobleaching and recovery. Biophys J. 1989 Sep;56(3):559–564. doi: 10.1016/S0006-3495(89)82703-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Meyer T. E., Yakali E., Cusanovich M. A., Tollin G. Properties of a water-soluble, yellow protein isolated from a halophilic phototrophic bacterium that has photochemical activity analogous to sensory rhodopsin. Biochemistry. 1987 Jan 27;26(2):418–423. doi: 10.1021/bi00376a012. [DOI] [PubMed] [Google Scholar]
  15. Monaco H. L., Zanotti G., Spadon P., Bolognesi M., Sawyer L., Eliopoulos E. E. Crystal structure of the trigonal form of bovine beta-lactoglobulin and of its complex with retinol at 2.5 A resolution. J Mol Biol. 1987 Oct 20;197(4):695–706. doi: 10.1016/0022-2836(87)90476-1. [DOI] [PubMed] [Google Scholar]
  16. Newcomer M. E., Jones T. A., Aqvist J., Sundelin J., Eriksson U., Rask L., Peterson P. A. The three-dimensional structure of retinol-binding protein. EMBO J. 1984 Jul;3(7):1451–1454. doi: 10.1002/j.1460-2075.1984.tb01995.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Nielsen J. B., Lampen J. O. Membrane-bound penicillinases in Gram-positive bacteria. J Biol Chem. 1982 Apr 25;257(8):4490–4495. [PubMed] [Google Scholar]
  18. Pervaiz S., Brew K. Homology and structure-function correlations between alpha 1-acid glycoprotein and serum retinol-binding protein and its relatives. FASEB J. 1987 Sep;1(3):209–214. doi: 10.1096/fasebj.1.3.3622999. [DOI] [PubMed] [Google Scholar]
  19. Scapin G., Gordon J. I., Sacchettini J. C. Refinement of the structure of recombinant rat intestinal fatty acid-binding apoprotein at 1.2-A resolution. J Biol Chem. 1992 Feb 25;267(6):4253–4269. doi: 10.2210/pdb1ifc/pdb. [DOI] [PubMed] [Google Scholar]
  20. Sundelin J., Das S. R., Eriksson U., Rask L., Peterson P. A. The primary structure of bovine cellular retinoic acid-binding protein. J Biol Chem. 1985 May 25;260(10):6494–6499. [PubMed] [Google Scholar]

Articles from Protein Science : A Publication of the Protein Society are provided here courtesy of The Protein Society

RESOURCES