A Rehmannia glutinosa caffeic acid O-methyltransferase functional identification: Reconstitution of the ferulic acid biosynthetic pathway in Saccharomyces cerevisiae using Rehmannia glutinosa enzymes.

Rehmannia glutinosa produces many pharmacological natural components, including ferulic acid (FA) which is also an important precursor of some medicinal ingredients, so it is very significant to explore FA biosynthesis for enhancing the production of FA and its derivations. This study aimed to determine and reconstitute the R. glutinosa FA biosynthetic pathway from phenylalanine (Phe) metabolism in Saccharomyces cerevisiae as a safe host for the biosynthesis of plant-derived products. Although plant caffeic acid O-methyltransferases (COMTs) are thought to be a vital catalytic enzyme in FA biosynthesis pathways, to date, none of the RgCOMTs in R. glutinosa has been characterized. This study identified an RgCOMT and revealed its protein enzymatic activity for FA production in vitro. The RgCOMT overexpression in R. glutinosa significantly increased FA yield, suggesting that its molecular function is involved in FA biosynthesis. Heterologous expression of the RgCOMT and reported R. glutinosa genes, RgPAL2 (encoding phenylalanine ammonia-lyase [PAL] protein), RgC4H (cinnamate 4-hydroxylase [C4H]), and RgC3H (p-coumarate-3-hydroxylase [C3H]), in S. cerevisiae confirmed their catalytic abilities in the reaction steps for the FA biosynthesis. Importantly, in this study, these genes were introduced into S. cerevisiae and coexpressed to reconstitute the R. glutinosa FA biosynthetic pathway from Phe metabolism, thus obtaining an engineered strain that produced an FA titer of 148.34 mg L-1 . This study identified the functional activity of RgCOMT and clarified the R. glutinosa FA biosynthesis pathway in S. cerevisiae, paving the way for the efficient production of FA and its derivatives.

[Effects of warming on physicochemical property of Cunninghamia lanceolata branch and leaf litter in subtropical plantation]. / 增温对亚热带杉木枝和叶凋落物理化性质的影响.

At the regional scale, substrate properties are the key factors driving litter decomposition rate. In this study, soil temperature was increased by buried heating cables to explore the impacts of climate warming on the physical and chemical properties in branch and leaf of Cunninghamia lanceolata litter. The results showed that after 5 years of soil warming (4 ℃), the contents of nitrogen (N), phosphorus (P) and water-soluble substance in branch litter increased by 35.2%, 40.8% and 7.6%, while that in leaf litter increased by 41.2%, 45.9% and 5.9%, respectively. The contents of carbon (C), cellulose and C/N in branch litter decreased by 5.1%, 11.6% and 28.8%, and in leaf litter decreased by 5.3%, 11.3% and 33.3%, respectively. Soil warming led to 29.8% increase in specific leaf area (SLA) and 40.7% decrease in tensile strength (LTS) of leaf litter. However, warming did not affect lignin content and pH value in both branch and leaf litter. 13C NMR and infrared spectrum analysis showed that the contents of amino acids, polysaccharides, polyphenols and aliphatic compounds in litter changed significantly after warming. Warming effect differed between litter organs, in that polysaccharides increased significantly only in leaf litter and the increase of amino acids in branch litter was greater than that in leaf litter. Overall, soil warming significantly changed the physical and chemical properties in C. lanceolata branch and leaf litter, which might accelerate the decomposition rate at the initial stage due to the increase of N, P contents and the decrease of LTS, but might decelerate the decomposition rate at the later stage due to an increase of complex polymers content in the litter.