Lembert N
Department of Histology and Cell Biology, Umeå University, Sweden.
Biochem J. 1996 Jul 1;317 ( Pt 1)(Pt 1):273-7. doi: 10.1042/bj3170273.
L-Luciferin is a competitive inhibitor of firefly luciferase with a K1 between 3 and 4 microM. Furthermore L-luciferin can serve as an alternative substrate for light production. Catalysis of L-luciferin can be observed in the absence of, or at low concentrations of, D-luciferin. The light production from L-luciferin increases slowly (maximal half-time 8 min) to a stable plateau. At low concentrations of enzyme and L-luciferin, maximal light production is about half of that observed at corresponding D-luciferin concentrations. Increasing the concentration of enzyme or L-luciferin reduces the light production relative to that obtained by D-luciferin catalysis. In contrast to the catalysis of D-luciferin the light production from L-luciferin can be effectively stimulated by the addition of PP1 provided that luciferase is premixed with inorganic pyrophosphatase (PP1-ase). A flash is emitted if PP1 is injected into a mixture of luciferase, L-luciferin, ATP and PP1-ase. The system maintains its responsiveness and emits further flashes of about equal duration and intensity upon repeated additions of PP1. It is proposed that PP1 induces a racemization of enzyme-bound L-luciferyl adenylate. The potential usefulness of PP1-dependent intracellular ATP monitoring is discussed. The proposed activation of firefly luciferase by PP1 may be part of the regulation of in vivo flashing.
L-荧光素是萤火虫荧光素酶的竞争性抑制剂，其K1在3至4微摩尔之间。此外，L-荧光素可作为发光的替代底物。在没有D-荧光素或D-荧光素浓度较低的情况下，可以观察到L-荧光素的催化作用。L-荧光素产生的光缓慢增加（最大半衰期8分钟）至稳定平台期。在低浓度的酶和L-荧光素条件下，最大光产量约为相应D-荧光素浓度下观察到的光产量的一半。相对于D-荧光素催化获得的光产量，增加酶或L-荧光素的浓度会降低光产量。与D-荧光素的催化作用相反，如果荧光素酶与无机焦磷酸酶（PP1酶）预混合，则添加PP1可有效刺激L-荧光素产生的光。如果将PP1注入荧光素酶、L-荧光素、ATP和PP1酶的混合物中，会发出闪光。该系统保持其响应性，并在重复添加PP1时发出持续时间和强度大致相等的进一步闪光。有人提出，PP1诱导酶结合的L-荧光素腺苷酸的消旋化。讨论了PP1依赖性细胞内ATP监测的潜在用途。所提出的PP1对萤火虫荧光素酶的激活可能是体内闪光调节的一部分。
