Separation of phycobiliprotein subunits by reverse-phase high-pressure liquid chromatography.
Baseline separation of subunits of diverse phycobiliproteins was achieved by a reverse-phase HPLC gradient method with a C4 large-pore column and a solvent system consisting of 0.1% trifluoroacetic acid (TFA) in water and 0.1% TFA in 2:1 (v/v) acetonitrile:isopropanol. The procedure was successfully applied to cyanobacterial allophycocyanin and C-phycocyanins, an unusual phycocyanin from a marine cyanobacterium, red algal B- and R-phycoerythrins, and a cryptomonad phycoerythrin. The subunit sizes in these proteins range from about 7.5 to 30 kDa. Sample recovery was in excess of 85% in all cases. On-line spectroscopic analysis with a multiple diode array detector allowed determination of the type and number of bilins carried by each subunit.
Swanson
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Division of Biochemistry and Molecular Biology, University of California, Berkeley 94720.
Glazer
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D000433:1-Propanol; D000097:Acetonitriles; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D045342:Light-Harvesting Protein Complexes; D010798:Phycocyanin / Q000302:isolation & purification; D010940:Plant Proteins / Q000302:isolation & purification*; D014269:Trifluoroacetic Acid
D000097:Acetonitriles; D045342:Light-Harvesting Protein Complexes; D010940:Plant Proteins; D010798:Phycocyanin; D000433:1-Propanol; D014269:Trifluoroacetic Acid