One-pot biotransformation of cellulose to amylose through multienzyme cascade with nucleoside diphosphate glucose as glycosyl donors

Artificial starch production from bioreactors is very promising in terms of amylose's broad applications as well as the possibility of addressing food shortage. We previously built an in vitro cellulose-to-starch pathway, synthesizing amylose from non-food cellulose. A challenge of this pathway lies in its low amylose yield due to the fact that only cellobiose in cellulose hydrolysate can be converted into amylose while cellodextrins with a degree of polymerization (DP) ≥ 3 cannot be utilized. Here, we report a new cellulose-to-amylose pathway through a multienzyme cascade where the phosphorolytic product glucose-1-phosphate (G-1-P) from both cellobiose and cellodextrins with DP ≥ 3 is converted to adenosine diphosphate glucose (ADPG) which then serves as a glycosyl donor for amylose synthesis. With introduction of a low-cost ATP regeneration system catalyzed by polyphosphate kinase (PPK), the amylose yield of one-pot biotransformation of cellulose reached 41.2 % at the cost of only 5 mM ATP input, exhibiting a 1.4-fold increase over the previous pathway. The new cellulose-to-amylose pathway has overcome the limitation in previous pathway and provided an efficient and sustainable bioprocess for artificial starch production as well as for utilization of agricultural residues.