Structural and biochemical approaches uncover multiple evolutionary trajectories of plant quinate dehydrogenases

Abstract

Quinate is produced and used by many plants in the biosynthesis of chlorogenic acids (CGAs). Chlorogenic acids are astringent and serve to deter herbivory. They also function as antifungal agents and have potent antioxidant properties. Quinate is produced at a branch point of shikimate biosynthesis by the enzyme quinate dehydrogenase (QDH). However, little information exists on the identity and biochemical properties of plant QDHs. In this study, we utilized structural and bioinformatics approaches to establish a QDH-specific primary sequence motif. Using this motif, we identified QDHs from diverse plants and confirmed their activity by recombinant protein production and kinetic assays. Through a detailed phylogenetic analysis, we show that plant QDHs arose directly from bifunctional dehydroquinate dehydratase-shikimate dehydrogenases (DHQD-SDHs) through different convergent evolutionary events, illustrated by our findings that eudicot and conifer QDHs arose early in vascular plant evolution whereas Brassicaceae QDHs emerged later. This process of recurrent evolution of QDH is further demonstrated by the fact that this family of proteins independently evolved NAD and NADP specificity in eudicots. The acquisition of QDH activity by these proteins was accompanied by the inactivation or functional evolution of the DHQD domain, as verified by enzyme activity assays and as reflected in the loss of key DHQD active site residues. The implications of QDH activity and evolution are discussed in terms of plant growth and development.

Evolution of a secondary metabolic pathway from primary metabolism: shikimate and quinate biosynthesis in plants

“…It appears plausible that following loss of QDH genes, some descending lineages faced selection pressures to regain QDH activity. This has indeed been found by Gritsunov et al (2018), who identified a recent SDH duplicate within the Brassicaceae that gained QDH independently.…”

“…Gritsunov et al . () identified a co‐factor shift from NADP to NAD within the angiosperm QDH clade and proposed an associated differentiation between catabolic and anabolic QDH functions within the angiosperm QDH clade. Furthermore, minute activity towards gallic acid biosynthesis has been detected in SDH enzymes from grape and walnut (Muir et al ., ; Bontpart et al ., ).…”

“…This has indeed been found by Gritsunov et al . (), who identified a recent SDH duplicate within the Brassicaceae that gained QDH independently.…”

“…Nevertheless, our data suggest that this mutation alone is sufficient to gain QDH activity. Likewise, introduction of the equivalent Thr to Gly change into the A. thaliana SDH protein is also sufficient to gain QDH activity (Gritsunov et al, 2018). In Arabidopsis, Lys385 and Asp423 have been identified as the major catalytic residues of the SDH active site (Singh and Christendat, 2006).…”

“…Within each clade, P. trichocarpa duplicates do not seem to be biochemically distinct, but show differential expression patterns during development and in response to environmental cues (Guo et al, 2014) suggesting physiological divergence. Gritsunov et al (2018) identified a co-factor shift from NADP to NAD within the angiosperm QDH clade and proposed an associated differentiation between catabolic and anabolic QDH functions within the angiosperm QDH clade. Furthermore, minute activity towards gallic acid biosynthesis has been detected in SDH enzymes from grape and walnut (Muir et al, 2011;Bontpart et al, 2016).…”