Agrobacterium sp. ATCC31750을 이용한 β-1,3-glucan 생산 공정에서 UDP-glucose 농도 변화

Abstract

Change of UDP-glucose concentration for β-1,3-glucan production using Agrobacterium sp. ATCC 31750

Park, Yang-Ho Advisor : Prof. Lee, Jung-Heon, Ph.D. Department of Chemical Engineering. Graduate School of Chosun University

Curdlan is one of biopolymer composed of β-1,3-glucan and dissolved in alkali solution but insoluble under neutral condition, and when the temperature of curdlan solution was heated over 80℃ it became gel. Agrobacterium sp. ATCC31750 grow in the presence of nitrogen source but produce β-1,3-glucan in the absence of the nitrogen source. Althought we tried to optimize system for the maximal production of β-1,3-glucan, the productivity of β-1,3-glucan still low. To increase β-1,3-glucan productivity, key metabolite UDP-glucose a precursor of β-1,3-glucan was selected and its concentration changes was measured. In this research, analysis of UDP-glucose by high performance liquid chromatography(HPLC) was established using a reversed phase system. The column used in this research was YMC-Pack Pro C18 column and 50 mM phosphate buffer solution was used as mobile phase. The flow rate of mobile phase was 1.0 ml/min, and UV wavelength was fixed at 254 nm. The change of UDP-glucose concentration was measured during cell growth and β-1,3-glucan production. The UDP-glucose concentration increases in cell growth level, UDP-glucose revelation activity is differed by a pH value in the Agrobacterium sp. fermentation and it was high in the environment of pH 5.5, also the β-1,3-glucan production showed high productivity. Using two-stage continuous fermentation process, the change of UDP-glucose concentration during β-1,3-glucan production period was measured. The pH of the first stage fermenter was controlled at pH 7.0, optimal for cell growth and that of 2nd stage was at pH 5.5, optimal for β-1,3-glucan production. When β-1,3-glucan production becomes steady state, the change of UDP-glucose concentration becomes steady state. Uracil was added to increase β-1,3-glucan production. The added uracil was used to activate UDP-glucose production, a precursor of β-1,3-glucan. When uracil was added at 84 hrs of fermentation the β-1,3-glucan productivity was decreased, the UDP-glucose concentration was suddenly increased and the β-1,3-glucan production was improved about 21 % more than that without uracil. Consequently, UDP-glucose played an important role for the synthesis of β-1,3-glucan. β-1,3-glucan production was increased under high concentration of UDP-glucose.