Fig. 5: Substrate promiscuity and subcellular localization of KDPG aldolase (EDA). | Nature Communications

Fig. 5: Substrate promiscuity and subcellular localization of KDPG aldolase (EDA).

From: Plastid ancestors lacked a complete Entner-Doudoroff pathway, limiting plants to glycolysis and the pentose phosphate pathway

Fig. 5

Chromatograms of reaction products GAP (a) and pyruvate (b) from an enzymatic assay of soybean and E. coli EDA when KDPG was supplied as substrate. GAP and pyruvate were analyzed by LCMS/MS and GC-MS, respectively. Authentic standards of GAP and pyruvate were used to confirm identities. Lineweaver-Burke plots for 2-keto-3-deoxy-6-phosphogluconate (KDPG) aldolase (EDA) activity are shown using KDPG (c) and D-fructose-1,6-bisphosphate (FBP) (d) as substrate at concentrations ranging from 0.25–1 mM (error bars signify standard deviation; n = 3). e Subcellular localization of GmEDA in agroinfiltrated Nicotiana benthamiana. Confocal images show green eGFP fluorescence, chlorophyll autofluorescence, and their overlays indicating GmEDA-eGFP colocalization with chloroplasts as punctate foci. Free eGFP and Arabidopsis thaliana 1-deoxy-D-xylulose 5-phosphate reductoisomerase (AtDXR) act as positive controls for cytosol and chloroplast localization, respectively. Bar = 50 μm. f Close-up of merged images for GmEDA and AtDXR showing colocalization detail of the eGFP fusion proteins with chloroplasts (bar = 5 μm). For (e) and (f), the experiment was performed twice with identical results.

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