Institut für Pharmazeutische Biologie und Biotechnologie, Philipps-Universität Marburg, Robert-Koch-Str. 4, 35037, Marburg, Germany.
Institut für Systematische und Evolutionäre Botanik, Universität Zürich, Zollikerstrasse 107, 8008, Zurich, Switzerland.
Planta. 2022 Jul 7;256(2):33. doi: 10.1007/s00425-022-03944-w.
Two isoforms of phenylalanine ammonia-lyase (PAL) have been isolated as cDNA sequences from the hornwort Anthoceros agrestis. The encoded enzymes convert L-phenylalanine and to lower extents L-tyrosine and L-histidine. Thus, the functional presence of the general phenylpropanoid pathway in one of the earliest land plant groups is established. The hornwort Anthoceros agrestis has an elaborated phenolic metabolism resulting in phenolic compounds, such as rosmarinic acid or megacerotonic acid. The general phenylpropanoid pathway is involved in the biosynthesis of these compounds. Two phenylalanine ammonia-lyase (PAL) genes, AaPAL1 and AaPAL2, have been identified in Anthoceros agrestis and the protein with an N-terminal 6xHis-tag heterologously synthesized in Escherichia coli for a full biochemical characterization. Both PAL proteins accept L-phenylalanine, L-tyrosine as well as L-histidine as substrates, although the activity is explicitly the highest with L-phenylalanine. K values as well as catalytic efficiencies were determined for phenylalanine (K AaPAL1 39 µM, AaPAL2 18 µM) and tyrosine (K AaPAL1 3.3 mM, AaPAL2 3.5 mM). In suspension cultures of Anthoceros agrestis, PAL genes were transcribed in parallel to rosmarinic acid (RA) accumulation and both showed highest abundance in the early growth phase. In a phylogenetic tree, both AaPAL amino acid sequences grouped within a clade with PAL amino acid sequences of diverse origin ranging from non-vascular to vascular plants, while most PALs from eudicots and monocots were mainly found in two other clades. The similarity of the hornwort PAL amino acid sequences to PAL sequences from vascular plants is more than 80% showing a strong conservation within the land plants. With this characterization of PALs from Anthoceros agrestis together with former investigations concerning cinnamic acid 4-hydroxylase and 4-coumaric acid CoA-ligase, the functional presence of the general phenylpropanoid pathway in this hornwort is proven.
已从角苔 Anthoceros agrestis 中分离出两种苯丙氨酸解氨酶（PAL）同工型的 cDNA 序列。编码的酶将 L-苯丙氨酸和低程度的 L-酪氨酸和 L-组氨酸转化。因此，在最早的陆地植物群之一中建立了一般苯丙烷途径的功能存在。角苔 Anthoceros agrestis 具有复杂的酚类代谢，导致产生酚类化合物，如迷迭香酸或megacerotonic 酸。一般苯丙烷途径参与这些化合物的生物合成。在 Anthoceros agrestis 中已经鉴定出两种苯丙氨酸解氨酶（PAL）基因，AaPAL1 和 AaPAL2，并在大肠杆菌中异源合成具有 N 端 6xHis 标签的蛋白质，以进行全面的生化特性分析。两种 PAL 蛋白都接受 L-苯丙氨酸、L-酪氨酸以及 L-组氨酸作为底物，尽管活性明显最高的是 L-苯丙氨酸。测定了苯丙氨酸（K AaPAL1 39µM，AaPAL2 18µM）和酪氨酸（K AaPAL1 3.3mM，AaPAL2 3.5mM）的 K 值和催化效率。在 Anthoceros agrestis 的悬浮培养物中，PAL 基因与迷迭香酸（RA）的积累平行转录，并且在早期生长阶段都表现出最高丰度。在系统发育树中，两种 AaPAL 氨基酸序列都属于与来自非维管束到维管束植物的各种来源的 PAL 氨基酸序列的一个分支，而大多数来自真双子叶植物和单子叶植物的 PAL 主要存在于另外两个分支中。角苔 PAL 氨基酸序列与维管束植物 PAL 序列的相似性超过 80%，在陆地植物中表现出很强的保守性。通过对角苔 Anthoceros agrestis 的 PAL 进行这种表征，以及以前关于肉桂酸 4-羟化酶和 4-香豆酸 CoA 连接酶的研究，证明了这种角苔中一般苯丙烷途径的功能存在。
