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Fig. 4 | Nutrition & Metabolism

Fig. 4

From: The interaction of selenoprotein F (SELENOF) with retinol dehydrogenase 11 (RDH11) implied a role of SELENOF in vitamin A metabolism

Fig. 4

The reductase activity of RDH11 toward all-trans-retinaldehyde affected by SELENOF. a.All-trans-retinol standard, it was eluted at 11.27 min; b. all-trans-retinal standard, it was eluted at 12.64 min; c. Homogenates (250 μl (concertiaon: 1 μg/μl) per reaction) of HEK293 T cells transfected with RDH11-YFP-N1 and HA-SelExpress1 blank vector; d. Homogenates (250 μl per reaction) of HEK293 T cells transfected with YFP-N1 blank vector and HA-SelenoF-SelExpress1; e. Homogenates (250 μl per reaction) of HEK293 T cells transfected with RDH11-YFP-N1 and HA-SELENOF-SelExpress1; f. Data analysis of all-trans-retinol produced in above c, d, e groups. The number 1, 2, 3 were corresponding to the group e, d, c respectively. The reaction products were quantified by summing the areas of both peaks of retinol and retinaldehyde. The ratios of the produced retinol were analyzed. The cell homogenates were incubated with 0.3 mmol/L all-trans-retinaldehyde in the presence of 0.3 mmol/L NADPH-Na4 for 15 min at 37 °C with 200 rpm shaking. The reactions were terminated by the addition of an equal volume of cold methanol: n-butyl alcohol 95:5 supplemented with 100 μg/ml butylated hydroxytoluene. Ten-microliter aliquots were analysed by reverse-phase HPLC using a C18 column. Mobile phase was acetonitrile: 30 mmol/L ammonium acetate (85: 15, v/v). The flow rate was 1.5 ml/min. Under these conditions, all-trans-retinol and all-trans-retinaldehyde eluted at around 11.27 and 12.64 min, respectively. Elution was monitored theabsorbance at 325 nm

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