Terpene synthases (TPSs) are pivotal enzymes for the production of diverse terpenes, including monoterpenes, sesquiterpenes, and diterpenes. In our recent studies, dictyostelid social amoebae, also known as cellular slime molds, were found to contain TPS genes for making volatile terpenes. For comparison, here we investigated Physarum polycephalum, a plasmodial slime mold also known as acellular amoeba. Plasmodia of P. polycephalum grown on agar plates were found to release a mixture of volatile terpenoids consisting of four major sesquiterpenes (α-muurolene, (E)-β-caryophyllene, two unidentified sesquiterpenoids) and the monoterpene linalool. There were no qualitative differences in terpenoid composition at two stages of young plasmodia. To understand terpene biosynthesis, we analyzed the transcriptome and genome sequences of P. polycephalum and identified four TPS genes designated PpolyTPS1-PpolyTPS4. They share 28-73% of sequence identities. Full-length cDNAs for the four TPS genes were cloned and expressed in Escherichia coli to produce recombinant proteins, which were tested for sesquiterpene synthase and monoterpene synthase activities. While neither PpolyTPS2 nor PpolyTPS3 was active, PpolyTPS1 and PpolyTPS4 were able to produce sesquiterpenes and monoterpenes from the respective substrates farnesyl diphosphate and geranyl diphosphate. By comparing the volatile profile of P. polycephalum plasmodia and the in vitro products of PpolyTPS1 and PpolyTPS4, it was concluded that most sesquiterpenoids emitted from P. polycephalum were attributed to PpolyTPS4. Phylogenetic analysis placed the four PpolyTPSs genes into two groups: PpolyTPS1 and PpolyTPS4 being one group that was clustered with the TPSs from the dictyostelid social amoeba and PpolyTPS2 and PpolyTPS3 being the other group that showed closer relatedness to bacterial TPSs. The biological role of the volatile terpenoids produced by the plasmodia of P. polycephalum is discussed.

中文翻译：

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疟原虫多头Phys的黏液中挥发性萜类化合物的排放和生物合成。

萜烯合酶（TPS）是生产各种萜烯（包括单萜烯，倍半萜烯和二萜烯）的关键酶。在我们最近的研究中，发现dictyostelid社交变形虫，也称为细胞粘液霉菌，含有用于制造挥发性萜烯的TPS基因。为了进行比较，在这里我们研究了Physarum polycephalum，一种浆状粘液霉菌，也被称为无细胞变形虫。发现在琼脂平板上生长的多头疟原虫的疟原虫释放出挥发性萜烯类化合物的混合物，该类萜类化合物由四种主要的倍半萜烯（α-茂丁烯，（E）-β-石竹烯，两个未鉴定的倍半萜类）和单萜芳樟醇组成。在年轻的疟原虫的两个阶段，萜类化合物的组成没有质的差异。为了了解萜烯的生物合成，我们分析了P的转录组和基因组序列。头cephal并鉴定了四个称为PpolyTPS1-PpolyTPS4的TPS基因。它们共享28-73％的序列同一性。克隆了四个TPS基因的全长cDNA，并在大肠杆菌中表达，以产生重组蛋白，并对其倍半萜合酶和单萜合酶活性进行了测试。尽管PpolyTPS2和PpolyTPS3均未激活，但PpolyTPS1和PpolyTPS4能够从相应的底物法呢基二磷酸酯和香叶基二磷酸酯生产倍半萜和单萜。通过比较多头疟原虫的浆状体和PpolyTPS1和PpolyTPS4的体外产物的挥发性特征，可以得出结论，多头疟原虫释放的大部分倍半萜类归因于PpolyTPS4。系统发育分析将四个PpolyTPSs基因分为两组：PpolyTPS1和PpolyTPS4是一组与双歧杆菌变形虫的TPS聚在一起的组，而PpolyTPS2和PpolyTPS3是另一组与细菌TPS密切相关的组。讨论了由多头疟原虫疟原虫产生的挥发性萜类化合物的生物学作用。