Enhanced curdlan production in Agrobacterium sp. ATCC 31749 by addition of low-polyphosphates

Abstract

A large amount of adenosine triphosphate with high energy phosphate bonds is required for uridine triphosphate regeneration during curdlan biosynthesis by Agrobacterium sp. ATCC 31749. To supply high energy for curdlan synthesis, three low-polyphosphates (Na4P2O7, Na5P3O10, and (NaPO3)6) with higher energy phosphate bonds were employed to substitute for KH2PO4-K2HPO4 in fermentation medium. Two genes encoding the polyphosphate metabolizing enzymes, polyphosphate kinase and exopolyphosphatase, were amplified and showed 95% homology to those in Agrobacterium sp. C58 by sequence analysis. The curdlan yields were enhanced by 23 and 134% when phosphate concentrations 0.024 mol/L of Na5P3O10 and 0.048 mol/L of (NaPO3)6 respectively, were added in the medium. The maximum curdlan yield of 30 ± 1.02 g/L was obtained with the addition of 0.048 mol/L of (NaPO3)6 with 5 g/L CaCO3 in the medium. When CaCO3 was removed from the culture and the three lowpolyphosphates were added, the pH and biomass yield dropped remarkably and little or no curdlan was produced. The culture containing 0.048 mol/L of (NaPO3)6 was mixed with KH2PO4-K2HPO4 and CaCO3 in the medium, but showed no effect on curdlan production. However, curdlan yield was improved by 49 ∼ 60% when CaCO3 was removed from the medium and KH2PO4-K2HPO4 acted as a buffer. It appears that the positive effect of (NaPO3)6 on curdlan production required the buffering capacity of CaCO3 and the absence of KH2PO4-K2HPO4 competing as a phosphate supplier.