Structural and morphological diversity of (1→3)-β-D-glucans synthesized in vitro by enzymes from Saprolegnia monoica.: Comparison with a corresponding in vitro product from blackberry (Rubus fruticosus)

Detergent extracts of microsomal fractions from Saprolegnia monoica and blackberry (Rubus fruticosus) cells were incubated with UDP-glucose to yield in vitro (1-->3)-beta-D-glucans. The insoluble products were analyzed by conventional and cryo transmission electron microscopy, X-ray diffraction, and C-13 CP/MAS NMR, and their molecular weights were determined by light scattering experiments. All the products were microfibrillar, but for the detergent extracts from S. monoica, important morphological differences were observed when the pH of the synthesizing medium was modified. At pH 6, the product had a weight average degree of polymerization ((DPw) over bar) exceeding 20 000 and consisted of endless ribbon-like microfibrils. The microfibrils obtained at pH 9 had a length of only 200-300 nm, and their (DPw) over bar was similar to5000. Of all the in vitro (1-->3)-beta-D-glucans, the one from R. fruticosus had the shortest length and the smallest (DPw) over bar. Crystallographic and spectroscopic data showed that the three in vitro samples consisted of triple helices of (1-->3)-beta-D-glucans and contained substantial amounts of water molecules in their structure, the shortest microfibrils being more hydrated. In addition, the long microfibrils from S. monoica synthesized at pH 6 were more resistant toward the action of an endo-(1-->3)-beta-D-glucanase than the shorter ones obtained at pH 9. These results are discussed in terms of molecular biosynthetic mechanisms of fungal and plant (1-->3)-beta-D-glucans, and in relation with the possible existence of several (1-->3)-beta-D-glucan synthases in a given organism. The interpretation and discussion of these observations integrate the current knowledge of the structure and function of (1-->3)-beta-D-glucans.