Location of starch granule-associated proteins revealed by confocal laser scanning microscopy

Distinct locations of starch granule-associated proteins were revealed using a protein-specific dye with confocal laser scanning microscopy (CLSM). The dye, 3-(4-carboxybenzoyl) quinoline-2-carboxaldchyde, fluoresces only after it reacts with primary amines in proteins, thereby removing background interference from residual dye. CLSM has the capability to discern fluorescence-labelled protein distribution in an optical slice of an intact starch granule while it is still in an intact state. With these techniques, starch granule proteins were revealed to be concentrated in internal concentric spheres in potato, maize, and wheat starches. Spheres were more distinct in potato starch than in other starches. Amylose-free potato and waxy maize starches showed no protein spheres, indicating that the internal protein spheres are composed of granule-bound starch synthase (GBSS). Identification of GBSS suggests (lie location of biosynthesis of amylose in starch granules, as well as spatial and temporal aspects of biosynthesis. (C) 2002 Elsevier Science Ltd.