Leont'eva O V, Vlasova T N, Ugarova N N
Faculty of Chemistry, Lomonosov Moscow State University, Moscow, Russia.
Biochemistry (Mosc). 2006 Jan;71(1):51-5. doi: 10.1134/s000629790601007x.
The absorption and fluorescence spectra of dimethyloxyluciferin (DMOL) and monomethyloxyluciferin (MMOL) were studied at pH 3.0-12.0. In the range of pH 3.0-8.0, the fluorescence spectrum of DMOL exhibits a maximum at lambda(em) = 639 nm. At higher pH values an additional emission maximum appears at lambda(em) = 500 nm (wavelength of excitation maximum lambda(ex) = 350 nm), which intensity increases with time. It is shown that this peak corresponds to the product of DMOL decomposition at pH > 8.0. The absorption spectra of MMOL were studied in the range of pH 6.0-9.0. At pH 8.0-9.0, the absorption spectrum of MMOL exhibits one peak at lambda(abs) = 440 nm. At pH 7.3-7.7, an additional band appears with maximum at lambda(abs) = 390 nm. At pH 6.0-7.0 two maxima are observed, at lambda(abs) = 375 and 440 nm. The fluorescence spectra of MMOL (pH 6.0-9.7, lambda(ex) = 440 or 375 nm) exhibit one maximum. It is shown that decomposition of DMOL and MMOL in aqueous solutions results in products of similar structure. DMOL and MMOL are rather stable at the pH optimum of luciferase. It is suggested that they can be used as fluorescent markers for investigation of the active site of the enzyme.
在pH 3.0 - 12.0条件下研究了二甲基氧化荧光素（DMOL）和单甲基氧化荧光素（MMOL）的吸收光谱和荧光光谱。在pH 3.0 - 8.0范围内，DMOL的荧光光谱在λ(em) = 639 nm处出现最大值。在较高pH值时，在λ(em) = 500 nm处出现另一个发射最大值（激发最大值波长λ(ex) = 350 nm），其强度随时间增加。结果表明，该峰对应于pH > 8.0时DMOL分解的产物。在pH 6.0 - 9.0范围内研究了MMOL的吸收光谱。在pH 8.0 - 9.0时，MMOL的吸收光谱在λ(abs) = 440 nm处呈现一个峰。在pH 7.3 - 7.7时，出现一个额外的吸收带，最大值在λ(abs) = 390 nm处。在pH 6.0 - 7.0时，观察到两个最大值，分别在λ(abs) = 375和440 nm处。MMOL的荧光光谱（pH 6.0 - 9.7，λ(ex) = 440或375 nm）呈现一个最大值。结果表明，DMOL和MMOL在水溶液中的分解产生结构相似的产物。DMOL和MMOL在荧光素酶的最适pH条件下相当稳定。建议它们可作为荧光标记物用于研究该酶的活性位点。
