Culture supernatant of bacterial species that utilize complex carbohydrates (e.g. Xylan, alginate, etc...) can present challenges when attempting to extract bacterial secreted proteins. I'll like to gather opinions on the most suitable approach to extract secreted protein from supernatant containing carbohydrates and other components that may interfere with 2D gels and/or shotgun proteomics.
I think the best way to extract the proteins would be with equal volume of tris saturated phenol (pH 8) followed by overnight precipitation (at -20C) with 100mM ammonium acetate in ice cold methanol.
Simplistic approach is size exclusion chromatography on something like sephacryl 200HR . Small broken down saccharides come out in the void (Vo) and your large complex in the the total volume (Vt) . This crude fractionation may help with a cleaner MALDI tof MS and partial fractionation of abundant common proteins from perhaps rare and interesting ones. Monitor elustion at 280 and 210nm use an ammonium bicarbonate buffer as when freeze drying your fractions to concentrate the buffer evaporates offer and you don't get high concentrations of salts in your test sample that often suppress ionisation of your target proteins.
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