title: Improvement in the thermostability of D-psicose 3-epimerase from Agrobacterium tumefaciens by random and site-directed mutagenesis.
abstract: The S213C, I33L, and I33L S213C variants of D-psicose 3-epimerase from Agrobacterium tumefaciens, which were obtained by random and site-directed mutagenesis, displayed increases of 2.5, 5, and 7.5°C in the temperature for maximal enzyme activity, increases of 3.3-, 7.2-, and 29.9-fold in the half-life at 50°C, and increases of 3.1, 4.3, and 7.6°C in apparent melting temperature, respectively, compared with the wild-type enzyme. Molecular modeling suggests that the improvement in thermostability in these variants may have resulted from increased putative hydrogen bonds and formation of new aromatic stacking interactions. The immobilized wild-type enzyme with and without borate maintained activity for 8 days at a conversion yield of 70% (350 g/liter psicose) and for 16 days at a conversion yield of 30% (150 g/liter psicose), respectively. After 8 or 16 days, the enzyme activity gradually decreased, and the conversion yields with and without borate were reduced to 22 and 9.6%, respectively, at 30 days. In contrast, the activities of the immobilized I33L S213C variant with and without borate did not decrease during the operation time of 30 days. These results suggest that the I33L S213C variant may be useful as an industrial producer of D-psicose.
authors.affiliation: Department of Bioscience and Biotechnology, Konkuk University, 1 Hwayang-Dong, Gwangjin-Gu, Seoul 143-701, Republic of Korea.
authors.forename: Jin-Geun
authors.initials: JG
authors.lastname: Choi
authors.forename: Yo-Han
authors.initials: YH
authors.lastname: Ju
authors.forename: Soo-Jin
authors.initials: SJ
authors.lastname: Yeom
authors.forename: Deok-Kun
authors.initials: DK
authors.lastname: Oh
mesh_terms: D016960:Agrobacterium tumefaciens / Q000201:enzymology* / Q000235:genetics; D019943:Amino Acid Substitution; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D005632:Fructose / Q000378:metabolism*; D008958:Models, Molecular; D016296:Mutagenesis; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D055550:Protein Stability; D019998:Racemases and Epimerases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D013696:Temperature; D044366:Transition Temperature
chemical_list: D004800:Enzymes, Immobilized; D050505:Mutant Proteins; C003243:psicose; D005632:Fructose; D019998:Racemases and Epimerases