AUTHOR=Zhao Haiying , Li Meng , Zhao Yuanyuan , Lin Xiaojing , Liang Huilin , Wei Jieshu , Wei Wuke , Ma Dongming , Zhou Zhongyu , Yang Jinfen 

TITLE=A Comparison of Two Monoterpenoid Synthases Reveals Molecular Mechanisms Associated With the Difference of Bioactive Monoterpenoids Between Amomum villosum and Amomum longiligulare

JOURNAL=Frontiers in Plant Science

VOLUME=12

YEAR=2021

URL=https://www.frontiersin.org/journals/plant-science/articles/10.3389/fpls.2021.695551

DOI=10.3389/fpls.2021.695551

ISSN=1664-462X

ABSTRACT=<p>The fruits of <italic>Amomum villosum</italic> and <italic>Amomum longiligulare</italic> are both used medicinally as <italic>Fructus Amomi</italic> the famous traditional Chinese medicine, however, the medicinal quality of <italic>A. villosum</italic> is better than that of <italic>A. longiligulare</italic>. Volatile terpenoids in the seeds, especially bornyl acetate and borneol, are the medicinal components of <italic>Fructus Amomi.</italic> The volatile terpenoids and transcriptome of developing seeds of <italic>A. villosum</italic> and <italic>A. longiligulare</italic> were compared in this study. The result revealed that the bornyl acetate and borneol contents were higher in <italic>A. villosum</italic> than in <italic>A. longiligulare</italic>. Additionally, six terpenoid synthase genes (<italic>AlTPS1–AlTPS6</italic>) were screened from the transcriptome of <italic>A. longiligulare</italic>, and <italic>AlTPS2</italic> and <italic>AlTPS3</italic> were found to share 98 and 83% identity with <italic>AvTPS2</italic> and <italic>AvBPPS</italic> (bornyl diphosphate synthase) from <italic>A. villosum</italic>, respectively. BPPS is the key enzyme for the biosynthesis of borneol and bornyl acetate. Biochemical assays improved that AlTPS2 had an identical function to AvTPS2 as linalool synthase; however, AlTPS3 produced camphene as the major product and bornyl diphosphate (BPP) as the secondary product, whereas AvBPPS produced BPP as its major product. There was only one different amino acid between AlTPS3 (A496) and AvBPPS (G495) in their conserved motifs, and the site-directed mutation of A496G in DTE motif of AlTPS3 changed the major product from camphene to BPP. Molecular docking suggests that A496G mutation narrows the camphene-binding pocket and decreases the BPP-binding energy, thus increases the product BPP selectivity of enzyme. In addition, the expression level of <italic>AvBPPS</italic> was significantly higher than that of <italic>AlTPS3</italic> in seeds, which was consistent with the related-metabolites contents. This study provides insight into the TPS-related molecular bases for the biosynthesis and accumulation differences of the bioactive terpenoids between <italic>A. villosum</italic> and <italic>A. longiligulare</italic>. <italic>BPPS</italic>, the key gene involved in the biosynthesis of the active compound, was identified as a target gene that could be applied for the quality-related identification and breeding of <italic>Fructus Amomi</italic>.</p>