Antimicrobial mechanism of theaflavins: They target 1-deoxy-D-xylulose 5-phosphate reductoisomerase, the key enzyme of the MEP terpenoid biosynthetic pathway

Abstract

1-Deoxy-D-xylulose 5-phosphate reductoisomerase (DXR) is the first committed enzyme in the 2-methyl-D-erythritol 4-phosphate (MEP) terpenoid biosynthetic pathway and is also a validated antimicrobial target. Theaflavins, which are polyphenolic compounds isolated from fermented tea, possess a wide range of pharmacological activities, especially an antibacterial effect, but little has been reported on their modes of antimicrobial action. To uncover the antibacterial mechanism of theaflavins and to seek new DXR inhibitors from natural sources, the DXR inhibitory activity of theaflavins were investigated in this study. The results show that all four theaflavin compounds could specifically suppress the activity of DXR, with theaflavin displaying the lowest effect against DXR (IC50 162.1 μM) and theaflavin-3,3′-digallate exhibiting the highest (IC50 14.9 μM). Moreover, determination of inhibition kinetics of the theaflavins demonstrates that they are non-competitive inhibitors of DXR against 1-deoxy-D-xylulose 5-phosphate (DXP) and un-competitive inhibitors with respect to NADPH. The possible interactions between DXR and the theaflavins were simulated via docking experiments.

Stability and stabilization of (–)‐gallocatechin gallate under various experimental conditions and analyses of its epimerization, auto‐oxidation, and degradation by LC‐MS

“…In fact, nitrogen, EDTA, and even superoxide dismutase (SOD) have been used to stabilize EGCG in PB and in HAM's F12‐RPMI 1640 medium, and nitrogen exhibited the best effect . We used VC as an antioxidant in our previous experiments to protect catechins and theaflavins . In this study, we employed nitrogen, EDTA, VC, and Na 2 SO 3 as protective reagents to stabilize GCG (20 μmol L −1 ) in the selected buffers (50 mmol L −1 , pH 7.4) and DMEM/F12 at 37 °C.…”

“…6 We used VC as an antioxidant in our previous experiments to protect catechins and theaflavins. [15][16][17] In this study, we wileyonlinelibrary.com/jsfa employed nitrogen, EDTA, VC, and Na 2 SO 3 as protective reagents to stabilize GCG (20 μmol L −1 ) in the selected buffers (50 mmol L −1 , pH 7.4) and DMEM/F12 at 37 ∘ C. Since some biological assays, particularly assays on cell level could need overnight or longer incubation, we performed the experiments in a 12 h period to better assess the stability of GCG under those conditions. First, EDTA at 10 mmol L −1 was tried, and the data depicted in Fig.…”

“…revealed that of all the tested tea catechins and theaflavins, GCG showed the strongest effect against Bacillus cereus . We recently investigated the antibacterial mechanisms of the tea polyphenols and found that the suppressive activity of GCG against 1‐deoxy‐ d ‐xylulose 5‐phosphate reductoisomerase is approximately ten times higher than that of EGCG . Therefore, it is highly necessary to fully elucidate the bioactivities of GCG.…”

Antimicrobial mechanism of epigallocatechin gallate and gallocatechin gallate: They target 1-deoxy-d-xylulose 5-phosphate reductoisomerase, the key enzyme of the MEP terpenoid biosynthetic pathway

“…In our previous study, we found that tea extracts exhibit moderate DXR inhibitory activity and the major active components seem to be polyphenols, especially EGCG and GCG [14]. We therefore further determined the details of DXR inhibition of the four theaflavin compounds [15] and the catechins EGCG and GCG so as to deepen our understanding of the antibacterial mechanism of these compounds and broaden our knowledge of tea. Furthermore, we could find through the study the DXR inhibitors of plant origin which possess completely different structural features compared to fosmidomycin.…”

“…The inhibitory activity of EGCG/GCG against E. coli DXR was determined by using the pre-column derivatization HPLC method published by this group with minor modification [15].…”