Figure 3.

Phosphorylation of Chlamydomonas starch occurs in C6 and C3 positions of the glucosyl residues. Following incubation of the algae with ³²P-orthophosphate in darkness for 2 h, starch was isolated and subjected to acid hydrolysis. An aliquot of the hydrolysate was mixed with authentic Glc6P and Glc3P, and a sample equivalent to 2,100 cpm was analyzed by HPAEC-PAD (top). Fractions were collected, and the radioactivity was determined (bottom). Fractions 2 and 4 comprise material coeluting with Glc6P and Glc3P, respectively. A total of 97% of the radioactivity applied to the column was recovered in fractions 1 to 5. Essentially the same result was obtained in an independent experiment (data not shown).