Zhang Liping, Li Haiyan, Gao Ling, Qi Yujie, Fu Wanyi, Li Xiongwei, Zhou Xiang, Gao Qikang, Gao Zhongshan, Jia Huijuan
Key Laboratory of Horticultural Plant Growth, Development and Quality improvement, Ministry of Agriculture, Department of Horticulture, Zhejiang University, Hangzhou, 310058, China.
Forest and Fruit Tree Institute, Shanghai Academy of Agricultural Sciences, Shanghai, 201403, China.
Plant Cell Rep. 2017 Jun;36(6):829-842. doi: 10.1007/s00299-017-2113-4. Epub 2017 Feb 25.
γ-Decalactone accumulation in peach mesocarp was highly correlated with ACX enzyme activity and natural PpACX1 content. Adding the purified recombinant PpACX1 induced γ-decalactone biosynthesis in cultured mesocarp discs in vitro. Previous gene expression studies have implied that acyl coenzyme A oxidase (ACX) is related to lactones synthesis, the characteristic aroma compounds of peach. Here, we analysed the correlation between γ-decalactone content and ACX enzyme activity in mesocarp of five different types of fully ripe peach varieties. Furthermore, 'Hu Jing Mi Lu' ('HJ') and 'Feng Hua Yu Lu' ('YL'), which have strong aroma among them, at four ripening stages were selected to study the role of ACX in lactone biosynthesis. The result showed that γ-decalactone was the most abundant lactone compound. γ-Decalactone accumulation was highly correlated with ACX enzyme activity. Mass spectrometry (MS) showed that PpACX1 was the most abundant PpACX protein in fully ripe mesocarp of cv. 'HJ'. To further elucidate the function of the PpACX1 protein, the PpACX1 gene was heterologously expressed in a bacterial system and characterized in vitro. MS identification gave the molecular weight of the recombinant PpACX1 as 94.44 kDa and the coverage rate of the peptide segments was 47.3%. In cultured mesocarp discs in vitro, adding the purified recombinant PpACX1 and C-CoA substrate induced the expected γ-decalactone biosynthesis. Using a sandwich ELISA based on mixed mono- and polyclonal antibodies against recombinant PpACX1, PpACX1 content in mesocarp was found to be highly correlated with γ-decalactone accumulation in mesocarp of five fully ripe varieties and four ripening stages of 'HJ' and 'YL'. This study revealed the vital function of PpACX1 in γ-decalactone biosynthesis in peach fruit.
桃中果皮中γ-癸内酯的积累与ACX酶活性和天然PpACX1含量高度相关。添加纯化的重组PpACX1可在体外培养的中果皮圆片中诱导γ-癸内酯的生物合成。先前的基因表达研究表明，酰基辅酶A氧化酶（ACX）与桃的特征香气化合物内酯的合成有关。在此，我们分析了五种不同类型的完全成熟桃品种中果皮中γ-癸内酯含量与ACX酶活性之间的相关性。此外，在其中具有强烈香气的‘湖景蜜露’（‘HJ’）和‘风华雨露’（‘YL’）的四个成熟阶段被选来研究ACX在内酯生物合成中的作用。结果表明，γ-癸内酯是最丰富的内酯化合物。γ-癸内酯的积累与ACX酶活性高度相关。质谱（MS）显示，PpACX1是‘HJ’品种完全成熟中果皮中最丰富的PpACX蛋白。为了进一步阐明PpACX1蛋白的功能，PpACX1基因在细菌系统中进行了异源表达并在体外进行了表征。MS鉴定给出重组PpACX1的分子量为94.44 kDa，肽段覆盖率为47.3%。在体外培养的中果皮圆片中，添加纯化的重组PpACX1和C-CoA底物可诱导预期的γ-癸内酯生物合成。使用基于针对重组PpACX1的混合单克隆和多克隆抗体的夹心ELISA，发现中果皮中的PpACX1含量与五个完全成熟品种以及‘HJ’和‘YL’的四个成熟阶段中果皮中γ-癸内酯的积累高度相关。本研究揭示了PpACX1在桃果实γ-癸内酯生物合成中的重要作用。
