Spicy SDS-PAGE gels: curcumin/turmeric as an environment-friendly protein stain

Abstract

Gel proteins are commonly stained with calorimetric/fluorescent dyes. Here, we demonstrate that heat-solubilized curcumin can serve as a nontoxic and environment-friendly fluorescent/colorimetric reversible protein stain. Curcumin, the yellow pigment found in the rhizomes of the perennial herb Curcuma longa (turmeric), is insoluble in aqueous solvents. However, heat (100°C) solubilization in water renders 1.5% of curcumin soluble. Curcumin solubilized by ethanol or alkali is ineffective in staining proteins. Heat solubilized curry spice turmeric stains proteins similarly. Staining is achieved in 30 min, with a sensitivity almost equaling that of Coomassie Brilliant Blue (CBB). Destaining is not required, and excess curcumin/turmeric can be discarded into the sink. Binding of proteins by silver inhibits curcumin binding, suggesting similarity of protein binding by silver and curcumin. It costs $1.5-2.0 to stain a mini-gel with curcumin, while turmeric costs less than 0.005 cent. CBB staining/destaining costs about 2 cents. However, CBB is toxic and its use necessitates specialized disposal efforts. Curcumin/turmeric, thus, can serve as an ideal nontoxic protein stain.

Fig. 1:

Staining of unstained protein molecular weight standards by heat solubilized curcumin and Coomassie Brilliant Blue (CBB). A- Stained by CBB; B- Stained by heat-solubilized curcumin for 30 min; C- Stained by heat-solubilized curcumin for 2 h. Lane 1 – 2 μg/protein marker; lane 2 – 1 μg/protein marker; lane 3 – 500 ng/protein marker; lane 4 – 100 ng/protein marker

Figure 2:

Comparison of protein staining efficacy of (A) curcumin solubilized in water with heat, (B) curcumin solubilized in absolute ethanol, (C) curcumin solubilized in DMSO (and diluted with water to obtain a 0.1% DMSO solution) and (D) curcumin solubilized in 0.5 N NaOH.

Figure 3:

Staining of various proteins by CBB and heat-solubilized curcumin or turmeric. IgM, Fc fragment, interleukin 13 receptor, mouse serum and bovine serum albumin stained with (A) CBB (B) heat solubilized curcumin (longer exposure) (C) heat solubilized curcumin (shorter exposure) (D) heat solubilized turmeric (E) stained with heat solubilized curcumin and visualized without ultraviolet light (F) stained with heat solubilized turmeric and visualized without ultraviolet light

Fig. 4:

Possible mechanism of curcumin (CU) binding to proteins. (Please note: Heat solubilized curcumin was used to stain BSA, whenever curcumin is mentioned) (A) Curcumin binding to BSA (B) Coomassie (CBB) binding to BSA (C) BSA stained first by heat solubilized curcumin, followed by staining with CBB (D) curcumin-stained BSA washed for two hours with methanol/acetic acid (25:10 % respectively) (E) BSA stained first with CBB, followed by staining with curcumin (F) BSA incubated first with silver (using standard silver staining protocol), followed by staining curcumin.