Enzyme-substrate complexes of the quinate/shikimate dehydrogenase from Corynebacterium glutamicum enable new insights in substrate and cofactor binding, specificity, and discrimination

Abstract

Quinate dehydrogenase (QDH) catalyzes the reversible oxidation of quinate to 3-dehydroquinate by nicotineamide adenine dinucleotide (NADH) and is involved in the catabolic quinate metabolism required for the degradation of lignin. The enzyme is a member of the family of shikimate/quinate dehydrogenases (SDH/QDH) occurring in bacteria and plants. We characterized the dual-substrate quinate/shikimate dehydrogenase (QSDH) from Corynebacterium glutamicum (CglQSDH) kinetically and revealed a clear substrate preference of CglQSDH for quinate compared with shikimate both at the pH optimum and in a physiological pH range, which is a remarkable contrast to closely related SDH/QDH enzymes. With respect to the cosubstrate, CglQSDH is strictly NAD(H) dependent. These substrate and cosubstrate profiles correlate well with the details of three atomic resolution crystal structures of CglQSDH in different functional states we report here: with bound NAD+ (binary complex) and as ternary complexes with NADH plus either shikimate or quinate. The CglQSDH-NADH-quinate structure is the first complex structure of any member of the SDH/QDH family with quinate. Based on this novel structural information and systematic sequence and structure comparisons with closely related enzymes, we can explain the strict NAD(H) dependency of CglQSDH as well as its discrimination between shikimate and quinate.

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

“…In contrast, SDHs show specificity for shikimate. Previous studies have used structural and mutational analyses to understand the basis of the broader substrate specificity of QDHs (Lindner et al ., ; Hoppner et al ., ). However, this approach has proved challenging, and so far attempts to understand the molecular basis for the different substrate preferences of the SDH and QDH classes have not been fruitful.…”

“…Quinate dehydrogenases have been more thoroughly studied in microbes, where they participate in the catabolism of quinate as a carbon source (Hoppner et al ., ). Microbial QDHs share a common structural fold with SDHs and catalyze the oxidation of both quinate and shikimate, with preference for quinate (Peek et al ., ; Hoppner et al ., ). In contrast, SDHs show specificity for shikimate.…”

De Novo Biosynthesis of Chlorogenic Acid Using an Artificial Microbial Community

“…Three dehydrogenases were compared. Among them, QSDH, 45 which has not been applied in QA production previously, exhibited the highest catalytic activity in synthesizing QA. In nature, a stable microbial community is essential for carrying out appropriate functions.…”