The enzymic utilization of sucrose in the synthesis of amylose and derivatives of amylose, using phosphorylases.

The synthesis of polysaccharides constitutes a challenge to organic chemists. This class of compounds is of current interest because of its importance in biological systemsl'2. Synthetic methods based on classical chemical manipulations are limited in applicabiiity: using these methods it is difficult to form glycosidic linkages stereospecifically, or to generate polymers of high molecular v,'eight3'a. Enzymic catalysis has proved to be an attractive alternative to classical chemical methods for the synthesis of polysacchandes:'5. In particular, (1+4)-a-o-glucan phosphorylase from potatoes (EC Z.a.L\ potato Jrhosphorylase; pph) catalyzes the reversible, degradative phosphorylation of anrllose to o-glucosyl phosphate (G-1-P), and has been used in the presence of primers to synthesize polvsaccharide chains5.6. This approach has two major disadvantages: the cost (-$1300/mol. Sigma) of the starting monomer G-1-P, and the-6G-80% yield of polymer based on this monomerT. We now report a new, coupled, enzymic system using sucrose phosphorylases (EC 2.4.1.7: sph) and pph that ,rtilizes inexpensive sucrose as the source of G-1-P. and gives >90Y" yields of high-molecular-weight amylose. using a variety of primers (see Fig. 1). In a representative experiment, sucrose (1: 1.77 g,5 mmol), maltoheptaose (a; 8 mg), and NaHzPOo Q4.5 mg; 5 mol%, relative to sucrose) were dissolved in 0.1rnl sodium citrate buffer (a0 mL) at 37'. The pH was adjusted to 7, and sph (Sigma,30 U) and pph5 (15 U) were added. Afterstirring for24 h, enzymic assaye for glucose, glucosyl phosphate, and fructose indicated that94"/" of the sucrose had been converted into amylose (5). The mixture was heated to 90o, the precipitate removed by filtration, and 1-butanol (a mL) was added to the supernatant liquor. After 24 h at 24" the precipitated amylose was filtered off , succcssively washed with water (10 mL), methanol (30 mL), and ether (50 mL), and dried in vacuo.