The investigation of violaxanthin de-epoxidation in the primitive green alga Mantoniella squamata (Prasinophyceae) indicates mechanistic differences in xanthophyll conversion to higher plants

摘要

The violaxanthin (Vx) de-epoxidation of the green alga Mantoniella squamata (Prasinophyceae) was investigated in thylakoid membranes and in enzyme assays with the isolated Vx de-epoxidase (VDE), where pure Vx or purified lightharvesting complexes of M. squamata (MLHC) or spinach (LHCII) were used. The detailed analysis of the VDE amino acid sequences of prasinophycean algae and higher plants suggests that structural differences of the M. squamata VDE are responsible for the slow conversion of Ax to Zx that is typical for this alga. The present experiments further demonstrate that the availability of the substrate Vx has a strong impact on the overall de-epoxidation reaction and the ratio of the second de-epoxidation rate (de-epoxidation from Ax to Zx) to the first de-epoxidation rate (conversion from Vx to Ax). High concentrations of available Vx, as found in enzyme assays with pure Vx, lead to saturation of the VDE and a strong competition with the intermediate reaction product Ax, thus decreasing the ratio of the second deepoxidation rate to the first de-epoxidation rate. In de-epoxidation assays with isolated LHCs, where the availability of Vx is limited by the detachment of Vx from the protein, an increased ratio of the second de-epoxidation rate to the first de-epoxidation rate is observed. The present study also shows that the interaction between the isolated LHC and the VDE can influence the ratio of the two de-epoxidation rates. Mantoniella squamata VDE, which exhibits a very low ratio of the second de-epoxidation rate to the first de-epoxidation rate in thylakoids or in enzyme assays with the MLHC, shows a high ratio when it is incubated with the LHCII of spinach.