The Protein-Chromophore Bond in B Phycoerythrin from Porphyridium cruentum. Radiosulfur Labeling Experiments

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Abstract
Red algae of the species P. cruentum were grown in a minimum sulfate medium containing 35SO42-. 35S-labeled phycoerythrin was extracted. B phycoerythrin, b phycoerythrin and R phycocyanin [light harvesting accessory pigments of photosynthesis] could be separated from other proteins by using a carrier-free electrophoresis on columns. The final ratio A545/A280 of B phycoerythrin obtained was .gtoreq. 5. 35S-labeled B phycoerythrin was digested proteolytically with trypsin and pepsin. The resulting 35S-containing bilipeptides were separated by isoelectric focusing. Zones of enhanced chromophore concentration always showed an enhanced radioactivity. Peptide fractions with a low molar ratio sulfur/chromophore (1.1-1.8) were purified to remove sucrose and the carrier ampholyte. A modified, optimized Edman degradation followed. A butylacetate-soluble, red Edman product was obtained that contained most of the chromophore and the bulk of the radioactivity. This product was purified by 2-dimensional TLC. The main spot of the chromatogram was subjected to acidic hydrolysis. The major part of the radioactivity in the hydrolysate cochromatographed with cysteine. That proves cysteine to be the binding amino acid in all cases investigated.