title: Refolding of firefly luciferase immobilized on agarose beads.
abstract: The renaturation yield of the denatured firefly luciferase decreased strongly with increasing protein concentration in a renaturation buffer, because of aggregation. In this study, firefly luciferase was immobilized on agarose beads at a high concentration. Although the protein concentration was extremely high (about 100-fold) compared to that of soluble luciferase, the renaturation yield was comparable with that for the soluble one. Thus, immobilization was shown to be effective for avoiding aggregation of firefly luciferase. It was also shown that the optimum buffer conditions for renaturation of the immobilized luciferase were the same as those for the renaturation in solution. Also, it was indicated that electrostatic interactions between a protein and the matrix have a negative effect on renaturation of the immobilized luciferase since the renaturation yield decreased at acidic pH only for the immobilized luciferase. These novel observations are described in detail in this paper.
authors.affiliation: Department of Chemistry and Biotechnology, Graduate School of Engineering, The University of Tokyo, Hongo, Bunkyo-ku, Tokyo 113-8656, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Zako
authors.forename: H
authors.initials: H
authors.lastname: Deguchi
authors.forename: A
authors.initials: A
authors.lastname: Kitayama
authors.forename: H
authors.initials: H
authors.lastname: Ueda
authors.forename: T
authors.initials: T
authors.lastname: Nagamune
mesh_terms: D000818:Animals; D001517:Coleoptera / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D008156:Luciferases / Q000737:chemistry*; D010710:Phosphates / Q000737:chemistry; D017510:Protein Folding; D020673:Protein Renaturation; D012685:Sepharose; D012965:Sodium Chloride / Q000737:chemistry; D013997:Time Factors
chemical_list: D010710:Phosphates; D012965:Sodium Chloride; D012685:Sepharose; D008156:Luciferases