Abstract
Properties of a recombinant aequorin were investigated in comparison with those of natural aequorin. In chromatographic behaviour the recombinant aequorin did not match any of ten isoaequorins tested, although it was very similar to aequorin J. Its sensitivity to Ca2+ was found to be higher than that of any isoaequorin except aequorin D. The recombinant aequorin exhibited no toxicity when tested in various kinds of cells, even where samples of natural aequorin had been found to be toxic. Properties of four recombinant semi-synthetic aequorins (fch-, hcp-, e- and n-types), prepared from the recombinant apo-aequorin and synthetic analogues of coelenterazine, were approximately parallel with those of corresponding semi-synthetic aequorins prepared from natural apo-aequorin. Both recombinant e-aequorin and natural e-aequorin J luminesced with high values of the luminescence intensity ratio I400/I465, although the ratios were not pCa-dependent. The recombinant aequorin and recombinant semi-synthetic aequorins are highly suited for monitoring cellular Ca2+.
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- Anctil M., Shimomura O. Mechanism of photoinactivation and re-activation in the bioluminescence system of the ctenophore Mnemiopsis. Biochem J. 1984 Jul 1;221(1):269–272. doi: 10.1042/bj2210269. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Blinks J. R., Wier W. G., Hess P., Prendergast F. G. Measurement of Ca2+ concentrations in living cells. Prog Biophys Mol Biol. 1982;40(1-2):1–114. doi: 10.1016/0079-6107(82)90011-6. [DOI] [PubMed] [Google Scholar]
- Fabiato A. Rapid ionic modifications during the aequorin-detected calcium transient in a skinned canine cardiac Purkinje cell. J Gen Physiol. 1985 Feb;85(2):189–246. doi: 10.1085/jgp.85.2.189. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Inouye S., Aoyama S., Miyata T., Tsuji F. I., Sakaki Y. Overexpression and purification of the recombinant Ca2+-binding protein, apoaequorin. J Biochem. 1989 Mar;105(3):473–477. doi: 10.1093/oxfordjournals.jbchem.a122689. [DOI] [PubMed] [Google Scholar]
- 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]
- 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]
- SHIMOMURA O., JOHNSON F. H., SAIGA Y. Extraction, purification and properties of aequorin, a bioluminescent protein from the luminous hydromedusan, Aequorea. J Cell Comp Physiol. 1962 Jun;59:223–239. doi: 10.1002/jcp.1030590302. [DOI] [PubMed] [Google Scholar]
- Shimomura O. Isolation and properties of various molecular forms of aequorin. Biochem J. 1986 Mar 1;234(2):271–277. doi: 10.1042/bj2340271. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Shimomura O., Johnson F. H. Properties of the bioluminescent protein aequorin. Biochemistry. 1969 Oct;8(10):3991–3997. doi: 10.1021/bi00838a015. [DOI] [PubMed] [Google Scholar]
- Shimomura O., Johnson F. H. Regeneration of the photoprotein aequorin. Nature. 1975 Jul 17;256(5514):236–238. doi: 10.1038/256236a0. [DOI] [PubMed] [Google Scholar]
- Shimomura O., Musicki B., Kishi Y. Semi-synthetic aequorin. An improved tool for the measurement of calcium ion concentration. Biochem J. 1988 Apr 15;251(2):405–410. doi: 10.1042/bj2510405. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Shimomura O., Musicki B., Kishi Y. Semi-synthetic aequorins with improved sensitivity to Ca2+ ions. Biochem J. 1989 Aug 1;261(3):913–920. doi: 10.1042/bj2610913. [DOI] [PMC free article] [PubMed] [Google Scholar]