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여주상,Jae Yeon Park,Sung Ho Yeom,유영제 한국생물공학회 2008 Biotechnology and Bioprocess Engineering Vol.13 No.1
Gluconate and glucose were selected as the carbon substrates in the production of poly-3-hydroxybutyrate (PHB). Gluconate was utilized to maximize the specific growth rate during the first stage of cell growth, whereas glucose was used to maximize PHB biosynthesis during the second stage of PHB accumulation. The sequential feeding of gluconate and glucose resulted in a 50% enhancement of PHB productivity as compared to the cultures cultivated on glucose alone. In conjunction with secondary glucose uptake, the presence of a trace amount of ammonium increased the rate of PHB biosynthesis during the stage of PHB accumulation. Via the feeding of 0.03 mmol/h of NH4Cl solution prior to the exhaustion of the initial amount of NH4Cl, PHB productivity was significantly enhanced as compared to the cultures raised on glucose alone. The glucose-grown culture evidenced a higher level of NADPH during the NH4Cl-exausted PHB accumulation stage than was observed in the gluconate-grown culture, which reflects that the reason of higher PHB production observed when glucose was used as a carbon source. NH4Cl feeding following the depletion of initial NH4Cl resulted in elevated levels of both ATP and NADPH, which increased the PHB biosynthesis rate, and also in a decrease in the level of NADH, which reflected the alleviation of the inhibitory effects on the cells caused by nitrogen depletion.
서정현,( Li Lin ),여주상,차형준 한국화학공학회 2007 화학공학의이론과응용 Vol.10 No.1
Baculoviral polyhedrin, which originated from Autographa califonica nuclear polyhedrosis virus, was employed for the first time as a novel fusion partner for expression of foreign proteins in Escherichia coli system. We characterized the expression of recombinant polyhedrin protein fused to green fluorescent protein (GFP). The polyhedrin fusion protein was successfully expressed as an insoluble inclusion body. The E. coli expressing polyhedrin-GFP fusion protein showed higher cell growth and higher GFP yield than the strain expressing soluble single GFP. Interestingly, the polyhedrin fusionportion showed almost the same characteristics as the native baculoviral polyhedrin; it was rapidly solubilized under alkaline conditions, similar to the conditions found in the insect midgut. In addition, the polyhedrin fusion portion was rapidly digested by alkaline proteases in insect Plutella xylostella midgut as well as by α-chymotrypsin, a protease that has similar properties to insect midgutpolyhedra-associated alkaline proteases. These unique properties suggest that baculoviral polyhedrin might be an advantageous fusion partner for production of foreign proteins, especially harmful proteins, in E. coli.