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. 1998 Sep;7(9):1915–1929. doi: 10.1002/pro.5560070907

Uclacyanins, stellacyanins, and plantacyanins are distinct subfamilies of phytocyanins: plant-specific mononuclear blue copper proteins.

A M Nersissian 1, C Immoos 1, M G Hill 1, P J Hart 1, G Williams 1, R G Herrmann 1, J S Valentine 1
PMCID: PMC2144163  PMID: 9761472

Abstract

The cDNAs encoding plantacyanin from spinach were isolated and characterized. In addition, four new cDNA sequences from Arabidopsis ESTs were identified that encode polypeptides resembling phytocyanins, plant-specific proteins constituting a distinct family of mononuclear blue copper proteins. One of them encodes plantacyanin from Arabidopsis, while three others, designated as uclacyanin 1, 2, and 3, encode protein precursors that are closely related to precursors of stellacyanins and a blue copper protein from pea pods. Comparative analyses with known phytocyanins allow further classification of these proteins into three distinct subfamilies designated as uclacyanins, stellacyanins, and plantacyanins. This specification is based on (1) their spectroscopic properties, (2) their glycosylation state, (3) the domain organization of their precursors, and (4) their copper-binding amino acids. The recombinant copper binding domain of Arabidopsis uclacyanin 1 was expressed, purified, and shown to bind a copper atom in a fashion known as "blue" or type 1. The mutant of cucumber stellacyanin in which the glutamine axial ligand was substituted by a methionine (Q99M) was purified and shown to possess spectroscopic properties similar to uclacyanin 1 rather than to plantacyanins. Its redox potential was determined by cyclic voltammetry to be +420 mV, a value that is significantly higher than that determined for the wild-type protein (+260 mV). The available structural data suggest that stellacyanins (and possibly other phytocyanins) might not be diffusible electron-transfer proteins participating in long-range electron-transfer processes. Conceivably, they are involved in redox reactions occurring during primary defense responses in plants and/or in lignin formation.

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Selected References

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  1. Adman E. T. Copper protein structures. Adv Protein Chem. 1991;42:145–197. doi: 10.1016/s0065-3233(08)60536-7. [DOI] [PubMed] [Google Scholar]
  2. Aikazyan V. Ts, Nalbandyan R. M. Studies on plantacyanin. I. Distribution in the plant kingdom, subcellular localization and physicochemical properties. Biochim Biophys Acta. 1981 Feb 27;667(2):421–432. doi: 10.1016/0005-2795(81)90208-7. [DOI] [PubMed] [Google Scholar]
  3. Altschul S. F., Gish W., Miller W., Myers E. W., Lipman D. J. Basic local alignment search tool. J Mol Biol. 1990 Oct 5;215(3):403–410. doi: 10.1016/S0022-2836(05)80360-2. [DOI] [PubMed] [Google Scholar]
  4. Antholine W. E., Hanna P. M., McMillin D. R. Low frequency EPR of Pseudomonas aeruginosa azurin. Analysis of ligand superhyperfine structure from a type 1 copper site. Biophys J. 1993 Jan;64(1):267–272. doi: 10.1016/S0006-3495(93)81363-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Baker E. N. Structure of azurin from Alcaligenes denitrificans refinement at 1.8 A resolution and comparison of the two crystallographically independent molecules. J Mol Biol. 1988 Oct 20;203(4):1071–1095. doi: 10.1016/0022-2836(88)90129-5. [DOI] [PubMed] [Google Scholar]
  6. Bao W., O'malley D. M., Whetten R., Sederoff R. R. A laccase associated with lignification in loblolly pine xylem. Science. 1993 Apr 30;260(5108):672–674. doi: 10.1126/science.260.5108.672. [DOI] [PubMed] [Google Scholar]
  7. Bergaman C., Gandvik E. K., Nyman P. O., Strid L. The amino acid sequence of Stellacyanin from the lacquer tree. Biochem Biophys Res Commun. 1977 Aug 8;77(3):1052–1059. doi: 10.1016/s0006-291x(77)80084-3. [DOI] [PubMed] [Google Scholar]
  8. Bradley D. J., Kjellbom P., Lamb C. J. Elicitor- and wound-induced oxidative cross-linking of a proline-rich plant cell wall protein: a novel, rapid defense response. Cell. 1992 Jul 10;70(1):21–30. doi: 10.1016/0092-8674(92)90530-p. [DOI] [PubMed] [Google Scholar]
  9. Chen C. G., Pu Z. Y., Moritz R. L., Simpson R. J., Bacic A., Clarke A. E., Mau S. L. Molecular cloning of a gene encoding an arabinogalactan-protein from pear (Pyrus communis) cell suspension culture. Proc Natl Acad Sci U S A. 1994 Oct 25;91(22):10305–10309. doi: 10.1073/pnas.91.22.10305. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Davin L. B., Wang H. B., Crowell A. L., Bedgar D. L., Martin D. M., Sarkanen S., Lewis N. G. Stereoselective bimolecular phenoxy radical coupling by an auxiliary (dirigent) protein without an active center. Science. 1997 Jan 17;275(5298):362–366. doi: 10.1126/science.275.5298.362. [DOI] [PubMed] [Google Scholar]
  11. Dennison C., Vijgenboom E., de Vries S., van der Oost J., Canters G. W. Introduction of a CuA site into the blue copper protein amicyanin from Thiobacillus versutus. FEBS Lett. 1995 May 22;365(1):92–94. doi: 10.1016/0014-5793(95)00429-d. [DOI] [PubMed] [Google Scholar]
  12. Du H., Simpson R. J., Moritz R. L., Clarke A. E., Bacic A. Isolation of the protein backbone of an arabinogalactan-protein from the styles of Nicotiana alata and characterization of a corresponding cDNA. Plant Cell. 1994 Nov;6(11):1643–1653. doi: 10.1105/tpc.6.11.1643. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Evans S. V. SETOR: hardware-lighted three-dimensional solid model representations of macromolecules. J Mol Graph. 1993 Jun;11(2):134-8, 127-8. doi: 10.1016/0263-7855(93)87009-t. [DOI] [PubMed] [Google Scholar]
  14. Fischer D., Eisenberg D. Protein fold recognition using sequence-derived predictions. Protein Sci. 1996 May;5(5):947–955. doi: 10.1002/pro.5560050516. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Guss J. M., Freeman H. C. Structure of oxidized poplar plastocyanin at 1.6 A resolution. J Mol Biol. 1983 Sep 15;169(2):521–563. doi: 10.1016/s0022-2836(83)80064-3. [DOI] [PubMed] [Google Scholar]
  16. Guss J. M., Merritt E. A., Phizackerley R. P., Freeman H. C. The structure of a phytocyanin, the basic blue protein from cucumber, refined at 1.8 A resolution. J Mol Biol. 1996 Oct 11;262(5):686–705. doi: 10.1006/jmbi.1996.0545. [DOI] [PubMed] [Google Scholar]
  17. Guss J. M., Merritt E. A., Phizackerley R. P., Hedman B., Murata M., Hodgson K. O., Freeman H. C. Phase determination by multiple-wavelength x-ray diffraction: crystal structure of a basic "blue" copper protein from cucumbers. Science. 1988 Aug 12;241(4867):806–811. doi: 10.1126/science.3406739. [DOI] [PubMed] [Google Scholar]
  18. Hart P. J., Nersissian A. M., Herrmann R. G., Nalbandyan R. M., Valentine J. S., Eisenberg D. A missing link in cupredoxins: crystal structure of cucumber stellacyanin at 1.6 A resolution. Protein Sci. 1996 Nov;5(11):2175–2183. doi: 10.1002/pro.5560051104. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hay M., Richards J. H., Lu Y. Construction and characterization of an azurin analog for the purple copper site in cytochrome c oxidase. Proc Natl Acad Sci U S A. 1996 Jan 9;93(1):461–464. doi: 10.1073/pnas.93.1.461. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Hibino T., Lee B. H., Takabe T., Takabe T. Expression and characterization of Met92Gln mutant plastocyanin from Silene pratensis. J Biochem. 1995 Jan;117(1):101–106. doi: 10.1093/oxfordjournals.jbchem.a124693. [DOI] [PubMed] [Google Scholar]
  21. Hippler M., Reichert J., Sutter M., Zak E., Altschmied L., Schröer U., Herrmann R. G., Haehnel W. The plastocyanin binding domain of photosystem I. EMBO J. 1996 Dec 2;15(23):6374–6384. [PMC free article] [PubMed] [Google Scholar]
  22. Holm Richard H., Kennepohl Pierre, Solomon Edward I. Structural and Functional Aspects of Metal Sites in Biology. Chem Rev. 1996 Nov 7;96(7):2239–2314. doi: 10.1021/cr9500390. [DOI] [PubMed] [Google Scholar]
  23. Kiefer-Meyer M. C., Gomord V., O'Connell A., Halpin C., Faye L. Cloning and sequence analysis of laccase-encoding cDNA clones from tobacco. Gene. 1996 Oct 31;178(1-2):205–207. doi: 10.1016/0378-1119(96)00381-2. [DOI] [PubMed] [Google Scholar]
  24. Kieliszewski M. J., Lamport D. T. Extensin: repetitive motifs, functional sites, post-translational codes, and phylogeny. Plant J. 1994 Feb;5(2):157–172. doi: 10.1046/j.1365-313x.1994.05020157.x. [DOI] [PubMed] [Google Scholar]
  25. Lewis N. G., Yamamoto E. Lignin: occurrence, biogenesis and biodegradation. Annu Rev Plant Physiol Plant Mol Biol. 1990;41:455–496. doi: 10.1146/annurev.pp.41.060190.002323. [DOI] [PubMed] [Google Scholar]
  26. Li S. X., Showalter A. M. Cloning and developmental/stress-regulated expression of a gene encoding a tomato arabinogalactan protein. Plant Mol Biol. 1996 Nov;32(4):641–652. doi: 10.1007/BF00020205. [DOI] [PubMed] [Google Scholar]
  27. Loopstra C. A., Sederoff R. R. Xylem-specific gene expression in loblolly pine. Plant Mol Biol. 1995 Jan;27(2):277–291. doi: 10.1007/BF00020183. [DOI] [PubMed] [Google Scholar]
  28. Malmström B. G. Rack-induced bonding in blue-copper proteins. Eur J Biochem. 1994 Aug 1;223(3):711–718. doi: 10.1111/j.1432-1033.1994.tb19044.x. [DOI] [PubMed] [Google Scholar]
  29. Mann K., Eckerskorn C., Mehrabian Z., Nalbandyan R. M. The amino acid sequence of the spinach basic cupredoxin plantacyanin. Biochem Mol Biol Int. 1996 Nov;40(5):881–887. doi: 10.1080/15216549600201493. [DOI] [PubMed] [Google Scholar]
  30. Mann K., Schäfer W., Thoenes U., Messerschmidt A., Mehrabian Z., Nalbandyan R. The amino acid sequence of a type I copper protein with an unusual serine- and hydroxyproline-rich C-terminal domain isolated from cucumber peelings. FEBS Lett. 1992 Dec 21;314(3):220–223. doi: 10.1016/0014-5793(92)81475-2. [DOI] [PubMed] [Google Scholar]
  31. Murata M., Begg G. S., Lambrou F., Leslie B., Simpson R. J., Freeman H. C., Morgan F. J. Amino acid sequence of a basic blue protein from cucumber seedlings. Proc Natl Acad Sci U S A. 1982 Nov;79(21):6434–6437. doi: 10.1073/pnas.79.21.6434. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Neale A. D., Wahleithner J. A., Lund M., Bonnett H. T., Kelly A., Meeks-Wagner D. R., Peacock W. J., Dennis E. S. Chitinase, beta-1,3-glucanase, osmotin, and extensin are expressed in tobacco explants during flower formation. Plant Cell. 1990 Jul;2(7):673–684. doi: 10.1105/tpc.2.7.673. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Nersissian A. M., Mehrabian Z. B., Nalbandyan R. M., Hart P. J., Fraczkiewicz G., Czernuszewicz R. S., Bender C. J., Peisach J., Herrmann R. G., Valentine J. S. Cloning, expression, and spectroscopic characterization of Cucumis sativus stellacyanin in its nonglycosylated form. Protein Sci. 1996 Nov;5(11):2184–2192. doi: 10.1002/pro.5560051105. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Nersissian A. M., Melkonyan V. Z., Nalbandyan R. M. Studies on plantacyanin. IV. Reconstitution with Cu-thionein, oxidation by cytochrome oxidase and autooxidation in the presence of cardiolipin. Biochim Biophys Acta. 1991 Feb 15;1076(3):337–342. doi: 10.1016/0167-4838(91)90473-d. [DOI] [PubMed] [Google Scholar]
  35. Pace C. N., Vajdos F., Fee L., Grimsley G., Gray T. How to measure and predict the molar absorption coefficient of a protein. Protein Sci. 1995 Nov;4(11):2411–2423. doi: 10.1002/pro.5560041120. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Pascher T., Karlsson B. G., Nordling M., Malmström B. G., Vänngård T. Reduction potentials and their pH dependence in site-directed-mutant forms of azurin from Pseudomonas aeruginosa. Eur J Biochem. 1993 Mar 1;212(2):289–296. doi: 10.1111/j.1432-1033.1993.tb17661.x. [DOI] [PubMed] [Google Scholar]
  37. Peisach J., Levine W. G., Blumberg W. E. Structural properties of stellacyanin, a copper mucoprotein from Rhus vernicifera, the Japanese lac tree. J Biol Chem. 1967 Jun 25;242(12):2847–2858. [PubMed] [Google Scholar]
  38. Pelham H. R. The retention signal for soluble proteins of the endoplasmic reticulum. Trends Biochem Sci. 1990 Dec;15(12):483–486. doi: 10.1016/0968-0004(90)90303-s. [DOI] [PubMed] [Google Scholar]
  39. Pogson B. J., Davies C. Characterization of a cDNA encoding the protein moiety of a putative arabinogalactan protein from Lycopersicon esculentum. Plant Mol Biol. 1995 May;28(2):347–352. doi: 10.1007/BF00020254. [DOI] [PubMed] [Google Scholar]
  40. Richards K. D., Schott E. J., Sharma Y. K., Davis K. R., Gardner R. C. Aluminum induces oxidative stress genes in Arabidopsis thaliana. Plant Physiol. 1998 Jan;116(1):409–418. doi: 10.1104/pp.116.1.409. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Romero A., Hoitink C. W., Nar H., Huber R., Messerschmidt A., Canters G. W. X-ray analysis and spectroscopic characterization of M121Q azurin. A copper site model for stellacyanin. J Mol Biol. 1993 Feb 20;229(4):1007–1021. doi: 10.1006/jmbi.1993.1101. [DOI] [PubMed] [Google Scholar]
  42. Ryden L., Lundgren J. O. On the evolution of blue proteins. Biochimie. 1979;61(7):781–790. doi: 10.1016/s0300-9084(79)80272-2. [DOI] [PubMed] [Google Scholar]
  43. Rydén L. G., Hunt L. T. Evolution of protein complexity: the blue copper-containing oxidases and related proteins. J Mol Evol. 1993 Jan;36(1):41–66. doi: 10.1007/BF02407305. [DOI] [PubMed] [Google Scholar]
  44. Sakurai T. Preparation and spectroscopic studies of cobalt(II)-substituted cucumber basic blue protein "plantacyanin". Biochem Biophys Res Commun. 1986 Sep 30;139(3):961–966. doi: 10.1016/s0006-291x(86)80271-6. [DOI] [PubMed] [Google Scholar]
  45. Satow Y., Cohen G. H., Padlan E. A., Davies D. R. Phosphocholine binding immunoglobulin Fab McPC603. An X-ray diffraction study at 2.7 A. J Mol Biol. 1986 Aug 20;190(4):593–604. doi: 10.1016/0022-2836(86)90245-7. [DOI] [PubMed] [Google Scholar]
  46. Schininà M. E., Maritano S., Barra D., Mondovì B., Marchesini A. Mavicyanin, a stellacyanin-like protein from zucchini peelings: primary structure and comparison with other cupredoxins. Biochim Biophys Acta. 1996 Sep 13;1297(1):28–32. doi: 10.1016/0167-4838(96)00079-9. [DOI] [PubMed] [Google Scholar]
  47. Schnabelrauch L. S., Kieliszewski M., Upham B. L., Alizedeh H., Lamport D. T. Isolation of pl 4.6 extensin peroxidase from tomato cell suspension cultures and identification of Val-Tyr-Lys as putative intermolecular cross-link site. Plant J. 1996 Apr;9(4):477–489. doi: 10.1046/j.1365-313x.1996.09040477.x. [DOI] [PubMed] [Google Scholar]
  48. Studier F. W., Rosenberg A. H., Dunn J. J., Dubendorff J. W. Use of T7 RNA polymerase to direct expression of cloned genes. Methods Enzymol. 1990;185:60–89. doi: 10.1016/0076-6879(90)85008-c. [DOI] [PubMed] [Google Scholar]
  49. Tagliavacca L., Moon N., Dunham W. R., Kaufman R. J. Identification and functional requirement of Cu(I) and its ligands within coagulation factor VIII. J Biol Chem. 1997 Oct 24;272(43):27428–27434. doi: 10.1074/jbc.272.43.27428. [DOI] [PubMed] [Google Scholar]
  50. Van Gysel A., Van Montagu M., Inzé D. A negatively light-regulated gene from Arabidopsis thaliana encodes a protein showing high similarity to blue copper-binding proteins. Gene. 1993 Dec 22;136(1-2):79–85. doi: 10.1016/0378-1119(93)90450-h. [DOI] [PubMed] [Google Scholar]
  51. Whetten R., Sederoff R. Lignin Biosynthesis. Plant Cell. 1995 Jul;7(7):1001–1013. doi: 10.1105/tpc.7.7.1001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Williams R. J. Energised (entatic) states of groups and of secondary structures in proteins and metalloproteins. Eur J Biochem. 1995 Dec 1;234(2):363–381. doi: 10.1111/j.1432-1033.1995.363_b.x. [DOI] [PubMed] [Google Scholar]
  53. Xu F., Shin W., Brown S. H., Wahleithner J. A., Sundaram U. M., Solomon E. I. A study of a series of recombinant fungal laccases and bilirubin oxidase that exhibit significant differences in redox potential, substrate specificity, and stability. Biochim Biophys Acta. 1996 Feb 8;1292(2):303–311. doi: 10.1016/0167-4838(95)00210-3. [DOI] [PubMed] [Google Scholar]
  54. van Driessche G., Dennison C., Sykes A. G., van Beeumen J. Heterogeneity of the covalent structure of the blue copper protein umecyanin from horseradish roots. Protein Sci. 1995 Feb;4(2):209–227. doi: 10.1002/pro.5560040208. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. von Heijne G. A new method for predicting signal sequence cleavage sites. Nucleic Acids Res. 1986 Jun 11;14(11):4683–4690. doi: 10.1093/nar/14.11.4683. [DOI] [PMC free article] [PubMed] [Google Scholar]

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