费氏发光杆菌荧光素酶的研究。IX. 酶 - 黄素单核苷酸中间体光谱特性的进一步研究。

Studies on luciferase from Photobacterium phosphoreum. IX. Further studies on the spectroscopic characteristics of the enzyme-FMN intermediates.

作者信息

Ashizawa N, Nakamura T, Watanabe T

出版信息

J Biochem. 1977 Apr;81(4):1057-62. doi: 10.1093/oxfordjournals.jbchem.a131528.

DOI:10.1093/oxfordjournals.jbchem.a131528

Abstract

The absorption and fluorometric changes of the reaction mixture of luciferase-FMNH2 complex with O2 were re-examined. Rapid formation (k2(app) = 2.0 s-1 at [O2] = 120 micrometer) of an intermediate with a single absorption maximum at 380 nm within the range of 350-550 nm, and a weak fluorescence at 520 nm (less than or equal to 10% of that of FMN when excited at 380 nm) was observed. The absorption and fluorescence spectra and decay rate of the intermediate were estimated by simulation using an analog computer. The decay rate (0.27 s-1 at 20 degrees C) was in agreement with that of an obligatory intermediate of the luminescent reaction previously determined by measuring aldehyde-initiated luminescence. The process of decay of X1 to FMN involved another intermediate X1' with spectroscopic characteristics rather similar to those of FMN.

摘要

对荧光素酶 - FMNH₂ 复合物与 O₂ 反应混合物的吸收和荧光变化进行了重新研究。在 350 - 550 nm 范围内，观察到在 380 nm 处有单一吸收最大值的中间体快速形成（在 [O₂] = 120 微米时，k2(app) = 2.0 s⁻¹），并且在 520 nm 处有微弱荧光（在 380 nm 激发时，小于或等于 FMN 荧光的 10%）。通过模拟使用模拟计算机估计中间体的吸收光谱、荧光光谱和衰减速率。衰减速率（在 20℃ 时为 0.27 s⁻¹）与先前通过测量醛引发的发光确定的发光反应的必需中间体的衰减速率一致。X1 衰减为 FMN 的过程涉及另一种中间体 X1'，其光谱特征与 FMN 相当相似。