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Carboxyl-terminal processing protease for the D1 precursor protein: cloning and sequencing of the spinach cDNA

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Abstract

A previous study has demonstrated that the carboxyl-terminal (C-terminal) processing protease in spinach for the D1 precursor protein (pD1) of the photosystem II reaction center is a monomeric protein of about 45 kDa. Based on the amino acid sequence data of the purified protease, a cDNA clone encoding the enzyme has been identified and sequenced, from a spinach green leaf cDNA library. In order to determine the 5′ end of the transcript, the rapid amplification of cDNA end (5′-RACE) technique was applied. By these analyses, the full-length transcript was established to consist of 1906 nucleotides and a poly(A) tail, containing an open reading frame (ORF) corresponding to a protein with 539 amino acid residues. By comparing the amino acid sequence of the purified protease with that deduced from nucleotide sequence of the cDNA clones, the enzyme was shown to be furnished with an extra amino-terminal extension characteristic of both a transit peptide and a signal sequence. This suggests that the protease is synthesized in the cytosol and translocated into the lumenal space of thylakoids. The mature part of the enzyme consists of 389 amino acid residues and exhibits a significant sequence homology with two groups of proteins as demonstrated by a computer homology search, i.e. (1) the deduced sequence of a protein proposed to be the C-terminal processing protease for pD1 in Synechocystis sp. PCC 6803, based on genetic experiments and (2) proteases for C-terminal cleavage identified in Escherichia coli and Bartonella bacilliformis.

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Abbreviations

DFP:

diisopropyl fluorophosphate

kDa:

kilodalton

NEM:

N-ethylmaleimide

ORF:

open reading frame

p-ABSF:

4-(2-aminoethyl)benzenesulfonyl fluoride

PAGE:

polyacrylamide gel electrophoresis

PCR:

polymerase chain reaction

pD1:

D1 precursor protein

PSII:

photosystem II

PVDF:

polyvinylidene difluoride

RACE:

rapid amplification of the cDNA end

RT-PCR:

reverse transcription-polymerase chain reaction

SDS:

sodium dodecyl sulfate

References

  1. Anbudurai PR, Mor TS, Ohad I, Shestakov SV, Pakrasi HB: The ctpA gene encodes the C-terminal processing protease for the D1 protein of the photosystem II reaction center complex. Proc Natl Acad Sci USA 91: 8082–8086 (1994).

    Google Scholar 

  2. Barrett AJ: Classification of peptidases. Meth Enzymol 244: 1–15 (1994).

    Google Scholar 

  3. Bowyer JR, Packer JCL, McCormack BA, Whitelegge JP, Robinson C, Taylor MA: Carboxyl-terminal processing of the D1 protein and photoactivation of water-splitting in photosystem II: partial purification and characterization of the processing enzyme from Scenedesmus obliquus and Pisum sativum. J Biol Chem 267: 5424–5433 (1992).

    Google Scholar 

  4. Chia CP, Arntzen CJ: Evidence for two-step processing of nuclear-encoded chloroplast proteins during membrane assembly. J Cell Biol 103: 725–731 (1986).

    Google Scholar 

  5. Cleveland DW, Fischer SG, Kirschner MW, Laemmli UK: Peptide mapping by limited proteolysis in sodium dodecyl sulfate and analysis by gel electrophoresis. J Biol Chem 252: 1102–1106 (1977).

    Google Scholar 

  6. Debus RJ, Barry BA, Babcock GT, McIntosh L: Site-directed mutagenesis identifies a tyrosine radical involved in the photosynthetic oxygen-evolving system. Proc Natl Acad Sci USA 85: 427–430 (1988).

    Google Scholar 

  7. Diner BA, Ries DF, Cohen BN, Metz JG: COOH-terminal processing of polypeptide D1 of the photosystem II reaction center of Scenedesmus obliquus is necessary for the assembly of the oxygen-evolving complex. J Biol Chem 263: 8972–8980 (1988).

    Google Scholar 

  8. Fujita S, Inagaki N, Yamamoto Y, Taguchi F, Matsumoto A, Satoh K: Identification of the carboxyl-terminal processing protease for the D1 precursor protein of the photosystem II reaction center of spinach. Plant Cell Physiol, 36: 1169–1177 (1995).

    Google Scholar 

  9. Gavel Y, von Heijne G: A conserved cleavage-site motif in chloroplast transit peptides. FEBS Lett 261: 455–458 (1990).

    Google Scholar 

  10. Hageman J, Robinson C, Smeekens S, Weisbeek P: A thylakoid processing protease is required for complete maturation of the lumenal protein plastocyanin. Nature 324: 567–569 (1986).

    Google Scholar 

  11. Hara H, Yamamoto Y, Higashitani A, Suzuki H, Nishimura Y: Cloning mapping, and characterization of the Escherichia coli prc gene, which is involved in C-terminal processing of penicillin-binding protein 3. J Bact 173: 4799–4813 (1991).

    Google Scholar 

  12. Inagaki N, Fujita S, Satoh K: Solubilization and partial purification of a thylakoidal enzyme of spinach involved in the processing of D1 protein. FEBS Lett 246: 218–222 (1989).

    Google Scholar 

  13. Inagaki N, Fujita S, Satoh K: Purification and properties of a thylakoidal enzyme of spinach involved in the processing of D1 protein of PS II reaction center. In: Baltscheffski M (eds) Current Research in Photosynthesis, pp. 763–766. Kluwer Academic Publishers. Dordrecht (1990).

    Google Scholar 

  14. Joshi CP: An inspection of the domain between putative TATA box and translation start site in 79 plant genes Nucl Acids Res 15: 6643–6653 (1987).

    Google Scholar 

  15. Kozak M: Compilation and analysis of sequences upstream from the translational start site in eukaryotic mRNAs. Nucl Acid Res 12: 857–872 (1984).

    Google Scholar 

  16. Laemmli UK: Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680–685 (1970).

    Google Scholar 

  17. Marder JB, Goloubinoff P, Edelman M: Molecular architecture of the rapidly metabolized 32-kilodalton protein of photosystem II: indications for COOH-terminal processing of a chloroplast membrane polypeptide. J Biol Chem 259: 3900–3908 (1984).

    Google Scholar 

  18. Metz JG, Seibert M: Presence in photosystem II core complexes of a 34-kilodalton polypeptide required for water photolysis. Plant Physiol 76: 829–832 (1984).

    Google Scholar 

  19. Minami E, Watanabe A: Detection of a precursor polypeptide of the rapidly-synthesized 32000-dalton thylakoid protein in spinach chloroplasts. Plant Cell Physiol 26: 839–846 (1985).

    Google Scholar 

  20. Nanba O, Satoh K: Isolation of a photosystem II reaction center consisting of D-1 and D-2 polypeptides and cytochrome b-559. Proc Natl Acad Sci USA 84: 109–112 (1987).

    Google Scholar 

  21. Nixon PJ, Trost JT, Diner BA: Role of the carboxy terminus of polypeptide D1 in the assembly of a functional water-oxidizing manganese cluster in photosystem II of the cyanobacterium Synechocystis sp. PCC 6803: assembly requires a free carboxyl group at C-terminal position 344. Biochemistry 31: 10859–10871 (1992).

    Google Scholar 

  22. Sambrook J, Fritsch EF, Maniatis T: Molecular Cloning: A Laboratory Manual, 2nd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY (1989).

    Google Scholar 

  23. Satoh K: Isolation and properties of the photosystem II reaction center. In: Deisenhofer J, Norris J (eds) The Photosynthetic Reaction Center, pp. 289–318. Academic Press, New York (1993).

    Google Scholar 

  24. Shackleton JB, Robinson C: Transport of proteins into chloroplasts: the thylakoidal processing peptidase is a signal-type peptidase with stringent substrate requirements at the −3 and −1 positions. J Biol Chem 266: 12152–12156 (1991).

    Google Scholar 

  25. Shestakov SV, Anbudurai PR, Stanbekova GE, Gadzhiev A, Lind LK, Pakrasi HB: Molecular cloning and characterization of the ctpA gene encoding a carboxyl-terminal processing protease: analysis of a spontaneous photosystem II-deficient mutant strain of the cyanobacterium Synechocystis sp. PCC 6803. J Biol Chem 269: 19354–19359 (1994).

    Google Scholar 

  26. Silber KR, Keiler KC, Sauer RT: Tsp: a tail-specific protease that selectively degrades proteins with nonpolar C termini. Proc Natl Acad Sci USA 89: 295–299 (1992).

    Google Scholar 

  27. Svensson B, Vass I, Styring S: Sequence analysis of the D1 and D2 reaction center proteins of photosystem II. Z Naturforsch 46c: 765–776 (1991).

    Google Scholar 

  28. Taguchi F, Yamamoto Y, Satoh K: Recognition of the structure around cleavage site by the carboxyl-terminal processing protease for D1 precursor protein of photosystem II reaction center. J Biol Chem 270: 10711–10716 (1995).

    Google Scholar 

  29. Takahashi M, Shiraishi T, Asada K: COOH-terminal residues of D1 and 44-kDa CPa-2 at spinach photosystem II core complex. FEBS Lett 240: 6–8 (1988).

    Google Scholar 

  30. Takahashi Y, Nakane H, Kojima H, Satoh K: Chromatographic purification and determination of the carboxy-terminal sequences of photosystem II reaction center protein, D1 and D2. Plant Cell Physiol 31: 273–280 (1990).

    Google Scholar 

  31. Taylor MA, Packer JCL, Bowyer JR: Processing of the D1 polypeptide of the photosystem II reaction center and photoactivation of a low fluorescence mutant (LF-1) of Scenedesmus obliquus. FEBS Lett 237: 229–233 (1988).

    Google Scholar 

  32. von Heijne G: Protein targeting signals. Curr Opin Cell Biol 2: 604–608 (1990).

    Google Scholar 

  33. Watanabe A, Price CA: Translation of mRNAs for subunits of chloroplast coupling factor 1 in spinach. Proc Natl Acad Sci USA 79: 6304–6308 (1982).

    Google Scholar 

  34. Zurawski G, Bohnert HJ, Whitfield PR, Bottomley W: Nucleotide sequence of the gene for the M r 32000 thylakoid membrane protein from Spinacia oleracea and Nicotiana debneyi predicts a totally conserved primary translation product of M r 38950. Proc Natl Acad Sci USA 79: 7699–7703 (1982).

    Google Scholar 

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Authors and Affiliations

  1. Division of Biological Regulation and Photobiology, National Institute for Basic Biology, 444, Okazaki, Japan

    Noritoshi Inagaki & Kimiyuki Satoh

  2. Department of Biology, Okayama University, 700, Okayama, Japan

    Yumiko Yamamoto & Kimiyuki Satoh

  3. School of Agricultural Science, Nagoya University, 464-01, Nagoya, Japan

    Hitoshi Mori

Authors
  1. Noritoshi Inagaki
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  2. Yumiko Yamamoto
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  3. Hitoshi Mori
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  4. Kimiyuki Satoh
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Inagaki, N., Yamamoto, Y., Mori, H. et al. Carboxyl-terminal processing protease for the D1 precursor protein: cloning and sequencing of the spinach cDNA. Plant Mol Biol 30, 39–50 (1996). https://doi.org/10.1007/BF00017801

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  • DOI: https://doi.org/10.1007/BF00017801

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