A unique homodimeric NAD(+)-linked isocitrate dehydrogenase from the smallest autotrophic eukaryote Ostreococcus tauri

作者：Tang, Wang-Gang; Song, Ping; Cao, Zheng-Yu; Wang, Peng; Zhu, Guo-Ping^*

来源：The FASEB Journal, 2015, 29(6): 2462-2472.

DOI：10.1096/fj.14-257014

• adaptation
• coenzyme specificity determinants
• kinetics
• phylogenetic analysis

In eukaryotes, NAD(+)-dependent isocitrate dehydrogenase (IDH) is strictly mitochondrial and is a key enzyme in the Krebs cycle. To date, all known NAD(+)-specific IDHs (NAD-IDHs) in the mitochondria are believed to be heteromeric in solution. Here, a unique homodimeric NAD-IDH from Ostreococcus tauri (OtIDH), the smallest autotrophic picoeukaryote, was unveiled. Active OtIDH has a molecular weight of similar to 93 kDa with each subunit of 46.7 kDa. In the presence of Mn2+ and Mg2+, OtIDH displayed 42-fold and 51-fold preference for NAD(+) over NADP(+), respectively. Interestingly, OtIDH exhibited a sigmoidal kinetic behavior in response to isocitrate unlike other homodimeric homologs, and a remarkably high affinity for isocitrate (S-0.5 < 10 mu M) unlike other heterooligomeric homologs. Furthermore, its coenzyme specificity can be completely converted from NAD(+) (ancient trait) to NADP(+) (adaptive trait) by rational mutagenesis based on the evolutionary trace. Mutants D344R and D344R/M345H displayed a 15-fold and 72-fold preference for NADP(+) over NAD(+), respectively, indicating that D344 and M345 are the determinants of NAD(+) specificity. These findings also suggest that OtIDH may be an ancestral form of type II IDHs (all reported members are NADP(+)-linked enzymes) and may have evolved into NADP(+)-dependent IDH for adaptation to the increased demand of NADPH under carbon starvation.

• 出版日期2015-6
• 单位安徽师范大学