Structure and composition of urate storage granules from the fat body of Manduca sexta

Abstract

Uric acid granules were isolated and purified from fat body of fifth instar tobacco hornworms, Manduca sexta (L.). Analysis by light microscopy indicated that preparations were homogeneous and all granules displayed birefringent properties. Scanning electron microscopy confirmed the homogeneity of the preparations and showed near spherical or biconcave shaped granules, many with hollow centers. The granules, 0.5–2.0 μm in dia, consisted of a tight array of concentrically whorled fibers about 0.02 μm in dia and 0.5–1.0 μm in length. Biochemical analyses revealed that the granules contained 75–78% uric acid, 4–5% protein and 0.5% carbohydrate. Elemental analysis indicated the presence of potassium (6.7%) with lesser amounts of sodium (0.5%) and that the granules contained 6–7% water. SDS-polyacrylamide gel electrophoresis indicated the presence of numerous peptides ranging in molecular weight from 20,000 to > 200,000. Amino acid analysis showed an abundance of aspartic acid, glutamic acid and apolar amino acids. Transmission electron microscopic analysis revealed that staining of individual fibers with uranyl acetate and lead citrate was restricted to the outer surfaces. These findings suggest that the granules are composed of tightly coiled fibers of crystalline uric acid/urates and that each fiber is enveloped with protein.