Catalytic Properties of Domain-Exchanged Chimeric Proteins between Firefly Luciferase and Drosophila Fatty Acyl-CoA Synthetase CG6178
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Firefly luciferase and fatty acyl-CoA synthetase are members of the acyl-CoA synthetase super family, which consists of a large N-terminal domain and a small C-terminal domain. Previously we found that firefly luciferase has fatty acyl-CoA synthetic activity, and also identified that the homolog of firefly luciferase in <I>Drosophila melanogaster</I> (CG6178) is a fatty acyl-CoA synthetase and is not a luciferase. In this study, we constructed chimeric proteins by exchanging the domain between <I>Photinus pyralis</I> luciferase (PpLase) and <I>Drosophila</I> CG6178, and determined luminescence and fatty acyl-CoA synthetic activities. A chimeric protein with the N-terminal domain of PpLase and the C-terminal domain of CG6178 (Pp/Dm) had luminescence activity, showing approximately 4% of the activity of wild-type luciferase. The Pp/Dm protein also had fatty acyl-CoA synthetic activity and the substrate specificity was similar to PpLase. In contrast, a chimeric protein with the N-terminal domain of CG6178 and the C-terminal of PpLase (Dm/Pp) had only fatty acyl-CoA synthetase activity, and the substrate specificity was similar to CG6178. These results suggest that the N-terminal domain of firefly luciferase is essential for substrate recognition, and that the C-terminal domain is indispensable but not specialized for the luminescence reaction.
- Agricultural and Biological Chemistry
Agricultural and Biological Chemistry 70(11), 2739-2744, 2006-11-23
Japan Society for Bioscience, Biotechnology, and Agrochemistry