Max Planck Institute for Chemical Ecology, Hans-Knöll-Strasse 8, D-07745, Jena, Germany.
Present Address: Michael Smith Laboratories, University of British Columbia, Vancouver, Canada.
BMC Plant Biol. 2018 Oct 22;18(1):251. doi: 10.1186/s12870-018-1478-z.
Nitrilases are nitrile-converting enzymes commonly found within the plant kingdom that play diverse roles in nitrile detoxification, nitrogen recycling, and phytohormone biosynthesis. Although nitrilases are present in all higher plants, little is known about their function in trees. Upon herbivory, poplars produce considerable amounts of toxic nitriles such as benzyl cyanide, 2-methylbutyronitrile, and 3-methylbutyronitrile. In addition, as byproduct of the ethylene biosynthetic pathway upregulated in many plant species after herbivory, toxic β-cyanoalanine may accumulate in damaged poplar leaves. In this work, we studied the nitrilase gene family in Populus trichocarpa and investigated the potential role of the nitrilase PtNIT1 in the catabolism of herbivore-induced nitriles.
A BLAST analysis revealed three putative nitrilase genes (PtNIT1, PtNIT2, PtNIT3) in the genome of P. trichocarpa. While PtNIT1 was expressed in poplar leaves and showed increased transcript accumulation after leaf herbivory, PtNIT2 and PtNIT3 appeared not to be expressed in undamaged or herbivore-damaged leaves. Recombinant PtNIT1 produced in Escherichia coli accepted biogenic nitriles such as β-cyanoalanine, benzyl cyanide, and indole-3-acetonitrile as substrates in vitro and converted them into the corresponding acids. In addition to this nitrilase activity, PtNIT1 showed nitrile hydratase activity towards β-cyanoalanine, resulting in the formation of the amino acid asparagine. The kinetic parameters of PtNIT1 suggest that the enzyme utilizes β-cyanoalanine and benzyl cyanide as substrates in vivo. Indeed, β-cyanoalanine and benzyl cyanide were found to accumulate in herbivore-damaged poplar leaves. The upregulation of ethylene biosynthesis genes after leaf herbivory indicates that herbivore-induced β-cyanoalanine accumulation is likely caused by ethylene formation.
Our data suggest a role for PtNIT1 in the catabolism of herbivore-induced β-cyanoalanine and benzyl cyanide in poplar leaves.
腈水解酶是植物界中常见的将腈转化为酶，在腈解毒、氮循环和植物激素生物合成中发挥着多样化的作用。尽管腈水解酶存在于所有高等植物中，但对其在树木中的功能知之甚少。在受到食草动物侵害后，杨树会产生大量的有毒腈，如苯甲腈、2-甲基丁腈和 3-甲基丁腈。此外，作为许多植物物种在受到食草动物侵害后上调的乙烯生物合成途径的副产物，有毒的β-氰基丙氨酸可能会在受损的杨树叶中积累。在这项工作中，我们研究了毛白杨中的腈水解酶基因家族，并研究了腈水解酶 PtNIT1 在食草动物诱导的腈代谢中的潜在作用。
BLAST 分析显示，在毛白杨基因组中存在三个假定的腈水解酶基因（PtNIT1、PtNIT2、PtNIT3）。虽然 PtNIT1 在杨树叶片中表达，并在叶片受到食草动物侵害后转录本积累增加，但 PtNIT2 和 PtNIT3 似乎在未受损或受食草动物侵害的叶片中不表达。在大肠杆菌中表达的重组 PtNIT1 可接受生物源性腈，如β-氰基丙氨酸、苯甲腈和吲哚-3-乙腈作为体外底物，并将其转化为相应的酸。除了这种腈水解酶活性外，PtNIT1 对β-氰基丙氨酸还表现出腈水解酶活性，导致天冬酰胺的形成。PtNIT1 的动力学参数表明，该酶在体内利用β-氰基丙氨酸和苯甲腈作为底物。事实上，β-氰基丙氨酸和苯甲腈在受食草动物侵害的杨树叶中积累。叶片受到食草动物侵害后，乙烯生物合成基因的上调表明，食草动物诱导的β-氰基丙氨酸积累可能是由乙烯形成引起的。
我们的数据表明，PtNIT1 在杨树叶片中食草动物诱导的β-氰基丙氨酸和苯甲腈的代谢中起作用。
