Synthesis of 4-methylvaleric acid, a precursor of pogostone, involves a 2-isobutylmalate synthase related to 2-isopropylmalate synthase of leucine biosynthesis

We show here that the side chain of pogostone, one of the major components of patchouli oil obtained from Pogostemon cablin and possessing a variety of pharmacological activities, is derived from 4-methylvaleric acid (4MVA). We also show that 4MVA is produced through the one-carbon α-ketoacid elongation (αKAE) pathway with the involvement of the key enzyme 2-isobutylmalate synthase (IBMS), a newly identified enzyme related to isopropylmalate synthase (IPMS) of Leu biosynthesis. Site-directed mutagenesis identified Met132 in the N-terminal catalytic region as affecting the substrate specificity of PcIBMS1. And even though PcIBMS1 possesses the C-terminal domain that in IPMS serves to mediate Leu inhibition, it is insensitive to Leu. The observation of the evolution of IBMS from IPMS, as well as previously reported examples of IPMS-related genes involved in making glucosinolates in Brassicaceae, acylsugars in Solanaceae, and flavor compounds in apple, indicate that IPMS genes represent an important pool for the independent evolution of genes for specialized metabolism.

Glucosinolate Biosynthesis: Role Of MAM Synthase And Its Perspectives

Glucosinolates, synthesized by the glucosinolate biosynthesis pathway, are the secondary metabolites used as a defence mechanism in the Brassicaceae plants, including Arabidopsis thaliana. The first committed step in the pathway, catalysed by methylthioalkylmalate (MAM) synthase (EC: 2.3.3.17), is to produce different variants of glucosinolates. Phylogenetic analyses suggest that possibly MAM synthases have been evolved from isopropylmalate synthase (IPMS) by the substitutions of five amino acid residues (L143I, H167L, S216G, N250G, and P252G) in the active site of IPMS due to point mutations. Considering the importance of MAM synthase in Brassicaceae plants, Petersen et al. (2019) made an effort to characterise the MAM synthase (15 MAM1 variants) in vitro by single substitution or double substitutions. In their study, the authors have expressed the variants in E. coli and analysed the amino acids in the cultures of E. coli in vivo. Since modifying the MAM synthases by transgenic approaches could increase the resistance of Brassicaceae plants for enhancing the defence effect of glucosinolates and their degraded products; hence, MAM synthases should be characterised in detail in vivo in A. thaliana along with the structural analysis of the enzyme for meaningful impact and for its imminent use in vivo.

Network And Meta-Omics Reveal The Key Degrader And Cooperation Patterns In An Efficient 1,4-Dioxane-Degrading Microbial Community

Background: 1,4-dioxane is an emerging wastewater contaminant with probable human carcinogenicity. Our current understanding of microbial interactions during 1,4-dioxane biodegradation process in mix cultures is limited. Here, we applied metagenomic, metatranscriptomic and co-occurrence network analyses to unraveling the microbial cooperation between degrader and non-degraders in an efficient 1,4-dioxane-degrading microbial community CH1.Results: The 1,4-dioxane degrading bacterium, Ancylobacter polymorphus ZM13, was isolated from CH1 and proved to be the key degrader because of the high relative abundance, highly expressed toluene monooxygenase genes tmoABCDEF and high betweenness centrality of networks. The strain ZM13 cooperated obviously with 6 bacterial genera in the network, among which Xanthobacter and Mesorhizobium were proved to be involved in the intermediate metabolism with responsible genes encoding alcohol dehydrogenase (adh), aldehyde dehydrogenase (aldh), glycolate oxidase (glcDEF), glyoxylate carboligase (gcl), malate synthase (glcB) and 2-isopropylmalate synthase (leuA) upregulated. Also, 1,4-dioxane facilitated the shift of biodiversity and function of CH1, and those cooperators of CH1 cooperated with ZM13 in the way of providing amino acids or fatty acids and relieving environmental stresses to promote biodegradation.Conclusions: This study revealed the biodiversity, community structure, microbial functions and interactions in a microbial community CH1 during the efficient 1,4-dioxane degradation and proved the degrader Ancylobacter polymorphus ZM13 that isolated from CH1 was the key degrading bacterium. These results provide new insights into our understanding of how the key degrading bacterium interacted with cooperators in a 1,4-dioxane-degrading community, and has important implications for predicting microbial cooperation and constructing highly efficient synthetic 1,4-dioxane-degrading communities.