Black pepper (Piper nigrum L.) is known for its high content of piperine, a cinnamoyl amide derivative regarded as largely responsible for the pungent taste of this widely used spice. Despite its long history and worldwide use, the biosynthesis of piperine and related amides has been enigmatic up to now. In this report we describe a specific piperic acid CoA ligase from immature green fruits of P. nigrum. The corresponding enzyme was cloned and functionally expressed in E. coli. The recombinant enzyme displays a high specificity for piperic acid and does not accept the structurally related feruperic acid characterized by a similar C-2 extension of the general C6-C3 phenylpropanoid structure. The enzyme is also inactive with the standard set of hydroxycinnamic acids tested including caffeic acid, 4-coumaric acid, ferulic acid, and sinapic acid. Substrate specificity is corroborated by in silico modelling that suggests a perfect fit for the substrate piperic acid to the active site of the piperic acid CoA ligase. The CoA ligase gene shows its highest expression levels in immature green fruits, is also expressed in leaves and flowers, but not in roots. Virus-induced gene silencing provided some preliminary indications that the production of piperoyl-CoA is required for the biosynthesis of piperine in black pepper fruits.

中文翻译：

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胡椒酸CoA连接酶可产生胡椒碱的假定前体，胡椒碱是黑胡椒果实的辛辣成分。

黑胡椒（Piper nigrum L.）以其高含量的胡椒碱而闻名，胡椒碱是一种肉桂酰基酰胺衍生物，被认为是造成这种广泛使用的香料辛辣味的主要原因。尽管胡椒碱及其相关酰胺的悠久历史和世界范围内的应用，但到目前为止，其生物合成一直是个谜。在此报告中，我们描述了来自黑假单胞菌未成熟绿色果实的一种特定的磷酸CoA连接酶。克隆了相应的酶并在大肠杆菌中功能性表达。重组酶对哌酸显示出高特异性，并且不接受与结构相关的阿魏酸，其特征在于一般C6-C3苯基丙酸结构的相似C-2延伸。该酶还与标准的一组羟肉桂酸无活性，包括咖啡酸，4-香豆酸，阿魏酸和芥子酸。通过计算机模拟证实了底物特异性，这表明底物胡椒酸与胡椒酸CoA连接酶的活性位点完美契合。CoA连接酶基因在未成熟的绿色水果中显示出最高的表达水平，在叶和花中也表达，但在根中却不表达。病毒诱导的基因沉默提供了一些初步的迹象，表明黑胡椒果实中胡椒碱的生物合成需要胡椒基辅酶A的产生。