Blue fluorescent protein from the calcium-sensitive photoprotein aequorin is a heat resistant enzyme, catalyzing the oxidation of coelenterazine.
Blue fluorescent protein from the calcium-sensitive photoprotein aequorin (BFP-aq) was prepared and determined to be a heat resistant enzyme, catalyzing the luminescent oxidation of coelenterazine (luciferin) with molecular oxygen as a general luciferase. After treatment with excess ethylenediaminetetraacetic acid to remove Ca2+ from BFP-aq, the blue fluorescence shifted to a greenish fluorescence. This greenish fluorescent protein (gFP-aq) was identified as a non-covalent complex of apoaequorin with coelenteramide (oxyluciferin) in a molar ratio of 1:1. By incubation with coelenterazine in the absence of reducing reagents, gFP-aq was converted to aequorin at 25 degrees C. BFP-aq and gFP-aq possessing both fluorescence and luminescence activities may work as novel reporter proteins.
Inouye
Satoshi
S
Yokohama Research Center, Chisso Corporation, 5-1 Okawa, Kanazawa-ku, Yokohama 236-8605, Japan. sinouye@chisso.co.jp
D000331:Aequorin / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D002384:Catalysis; D002942:Circular Dichroism; D006358:Hot Temperature; D007093:Imidazoles / Q000378:metabolism*; D007202:Indicators and Reagents; D008164:Luminescent Proteins / Q000302:isolation & purification / Q000378:metabolism*; D010084:Oxidation-Reduction; D011719:Pyrazines / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D007093:Imidazoles; D007202:Indicators and Reagents; D008164:Luminescent Proteins; D011719:Pyrazines; C409504:blue fluorescent protein, Aequorea victoria; C017144:coelenterazine; D000331:Aequorin; D002118:Calcium