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. 1989 Sep;86(17):6567–6571. doi: 10.1073/pnas.86.17.6567

Cloning and expression of cDNA for the luciferase from the marine ostracod Vargula hilgendorfii.

E M Thompson 1, S Nagata 1, F I Tsuji 1
PMCID: PMC297885  PMID: 2771943

Abstract

The marine ostracod Vargula hilgendorfii ejects luciferin and luciferase into seawater to produce a bright luminous cloud. The light is due to the oxidation of luciferin, an imidazopyrazine compound, by molecular oxygen, catalyzed by luciferase. The mechanism of the reaction has been studied extensively and the 60 kcal/mol required for the blue emission have been shown to be derived from the oxidation of luciferin via a dioxetanone intermediate, in which the excited state oxyluciferin bound to luciferase is the emitter. However, only limited information is available regarding the properties of the enzyme. This paper reports the cloning and sequence analysis of the cDNA for Vargula luciferase and the expression of the cDNA in a mammalian cell system. The primary structure, deduced from the nucleotide sequence, consists of 555 amino acid residues in a single polypeptide chain with a molecular weight of 62,171. Two regions of the enzyme show significant amino acid sequence homology with an N-terminal segment of the photoprotein aequorin. The Vargula luciferase gene, which contains a signal sequence for secretion, should be well suited as a reporter in studies of gene expression.

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Selected References

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

  1. Aviv H., Leder P. Purification of biologically active globin messenger RNA by chromatography on oligothymidylic acid-cellulose. Proc Natl Acad Sci U S A. 1972 Jun;69(6):1408–1412. doi: 10.1073/pnas.69.6.1408. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Benton W. D., Davis R. W. Screening lambdagt recombinant clones by hybridization to single plaques in situ. Science. 1977 Apr 8;196(4286):180–182. doi: 10.1126/science.322279. [DOI] [PubMed] [Google Scholar]
  3. Chen E. Y., Seeburg P. H. Supercoil sequencing: a fast and simple method for sequencing plasmid DNA. DNA. 1985 Apr;4(2):165–170. doi: 10.1089/dna.1985.4.165. [DOI] [PubMed] [Google Scholar]
  4. Chirgwin J. M., Przybyla A. E., MacDonald R. J., Rutter W. J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. doi: 10.1021/bi00591a005. [DOI] [PubMed] [Google Scholar]
  5. Engebrecht J., Simon M., Silverman M. Measuring gene expression with light. Science. 1985 Mar 15;227(4692):1345–1347. doi: 10.1126/science.2983423. [DOI] [PubMed] [Google Scholar]
  6. Gluzman Y. SV40-transformed simian cells support the replication of early SV40 mutants. Cell. 1981 Jan;23(1):175–182. doi: 10.1016/0092-8674(81)90282-8. [DOI] [PubMed] [Google Scholar]
  7. Gorman C. M., Moffat L. F., Howard B. H. Recombinant genomes which express chloramphenicol acetyltransferase in mammalian cells. Mol Cell Biol. 1982 Sep;2(9):1044–1051. doi: 10.1128/mcb.2.9.1044. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Graham F. L., van der Eb A. J. A new technique for the assay of infectivity of human adenovirus 5 DNA. Virology. 1973 Apr;52(2):456–467. doi: 10.1016/0042-6822(73)90341-3. [DOI] [PubMed] [Google Scholar]
  9. Hauber R., Miska W., Schleinkofer L., Geiger R. The application of a photon-counting camera in very sensitive, bioluminescence-enhanced detection systems for protein blotting. Ultrasensitive detection systems for protein blotting and DNA hybridization, II. J Clin Chem Clin Biochem. 1988 Mar;26(3):147–148. [PubMed] [Google Scholar]
  10. Inouye S., Noguchi M., Sakaki Y., Takagi Y., Miyata T., Iwanaga S., Miyata T., Tsuji F. I. Cloning and sequence analysis of cDNA for the luminescent protein aequorin. Proc Natl Acad Sci U S A. 1985 May;82(10):3154–3158. doi: 10.1073/pnas.82.10.3154. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. JOHNSON F. H., SUGIYAMA N., SHIMOMURA O., SAIGA Y., HANEDA Y. Crystalline luciferin from a luminescent fish, Parapriacanthus beryciformes. Proc Natl Acad Sci U S A. 1961 Apr 15;47:486–489. doi: 10.1073/pnas.47.4.486. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kozak M. How do eucaryotic ribosomes select initiation regions in messenger RNA? Cell. 1978 Dec;15(4):1109–1123. doi: 10.1016/0092-8674(78)90039-9. [DOI] [PubMed] [Google Scholar]
  13. Kurose K., Inouye S., Sakaki Y., Tsuji F. I. Bioluminescence of the Ca2+-binding photoprotein aequorin after cysteine modification. Proc Natl Acad Sci U S A. 1989 Jan;86(1):80–84. doi: 10.1073/pnas.86.1.80. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Lynch R. V., 3rd, Tsuji F. I., Donald D. H. Evidence for a calcium requirement in the Cypridina bioluminescence reaction. Biochem Biophys Res Commun. 1972 Feb 25;46(4):1544–1550. doi: 10.1016/0006-291x(72)90783-8. [DOI] [PubMed] [Google Scholar]
  15. Marshall R. D. Glycoproteins. Annu Rev Biochem. 1972;41:673–702. doi: 10.1146/annurev.bi.41.070172.003325. [DOI] [PubMed] [Google Scholar]
  16. Mitchell G. W., Hastings J. W. A stable, inexpensive, solid-state photomultiplier photometer. Anal Biochem. 1971 Jan;39(1):243–250. doi: 10.1016/0003-2697(71)90481-7. [DOI] [PubMed] [Google Scholar]
  17. Morishita K., Kubota N., Asano S., Kaziro Y., Nagata S. Molecular cloning and characterization of cDNA for human myeloperoxidase. J Biol Chem. 1987 Mar 15;262(8):3844–3851. [PubMed] [Google Scholar]
  18. Ohtsuka E., Matsuki S., Ikehara M., Takahashi Y., Matsubara K. An alternative approach to deoxyoligonucleotides as hybridization probes by insertion of deoxyinosine at ambiguous codon positions. J Biol Chem. 1985 Mar 10;260(5):2605–2608. [PubMed] [Google Scholar]
  19. Ow D. W., DE Wet J. R., Helinski D. R., Howell S. H., Wood K. V., Deluca M. Transient and stable expression of the firefly luciferase gene in plant cells and transgenic plants. Science. 1986 Nov 14;234(4778):856–859. doi: 10.1126/science.234.4778.856. [DOI] [PubMed] [Google Scholar]
  20. Prasher D. C., McCann R. O., Longiaru M., Cormier M. J. Sequence comparisons of complementary DNAs encoding aequorin isotypes. Biochemistry. 1987 Mar 10;26(5):1326–1332. doi: 10.1021/bi00379a019. [DOI] [PubMed] [Google Scholar]
  21. Prasher D., McCann R. O., Cormier M. J. Cloning and expression of the cDNA coding for aequorin, a bioluminescent calcium-binding protein. Biochem Biophys Res Commun. 1985 Feb 15;126(3):1259–1268. doi: 10.1016/0006-291x(85)90321-3. [DOI] [PubMed] [Google Scholar]
  22. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Shimomura O., Johnson F. H., Masugi T. Cypridina bioluminescence: light-emitting oxyluciferin-luciferase complex. Science. 1969 Jun 13;164(3885):1299–1300. doi: 10.1126/science.164.3885.1299. [DOI] [PubMed] [Google Scholar]
  24. Shimomura O., Johnson F. H. Mechanism of the luminescent oxidation of cypridina luciferin. Biochem Biophys Res Commun. 1971 Jul 16;44(2):340–346. doi: 10.1016/0006-291x(71)90605-x. [DOI] [PubMed] [Google Scholar]
  25. Stone H. The enzyme catalyzed oxidation of Cypridina luciferin. Biochem Biophys Res Commun. 1968 May 10;31(3):386–391. doi: 10.1016/0006-291x(68)90487-7. [DOI] [PubMed] [Google Scholar]
  26. Tsuji F. I., Haneda Y., Lynch R. V., 3rd, Sugiyama N. Luminescence cross-reactions of Porichthys luciferin and theories on the origin of luciferin in some shallow-water fishes. Comp Biochem Physiol A Comp Physiol. 1971 Sep 1;40(1):163–179. doi: 10.1016/0300-9629(71)90159-9. [DOI] [PubMed] [Google Scholar]
  27. Tsuji F. I., Lynch R. V., 3rd, Stevens C. L. Some properties of luciferase from the bioluminescent crustacean, Cypridina hilgendorfii. Biochemistry. 1974 Dec 3;13(25):5204–5209. doi: 10.1021/bi00722a024. [DOI] [PubMed] [Google Scholar]
  28. Wahl G. M., Stern M., Stark G. R. Efficient transfer of large DNA fragments from agarose gels to diazobenzyloxymethyl-paper and rapid hybridization by using dextran sulfate. Proc Natl Acad Sci U S A. 1979 Aug;76(8):3683–3687. doi: 10.1073/pnas.76.8.3683. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Wigler M., Pellicer A., Silverstein S., Axel R. Biochemical transfer of single-copy eucaryotic genes using total cellular DNA as donor. Cell. 1978 Jul;14(3):725–731. doi: 10.1016/0092-8674(78)90254-4. [DOI] [PubMed] [Google Scholar]
  30. de Wet J. R., Wood K. V., DeLuca M., Helinski D. R., Subramani S. Firefly luciferase gene: structure and expression in mammalian cells. Mol Cell Biol. 1987 Feb;7(2):725–737. doi: 10.1128/mcb.7.2.725. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. von Heijne G. Patterns of amino acids near signal-sequence cleavage sites. Eur J Biochem. 1983 Jun 1;133(1):17–21. doi: 10.1111/j.1432-1033.1983.tb07424.x. [DOI] [PubMed] [Google Scholar]

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