title: Association and activation of fructose 1,6-bisphosphase during unfolding and refolding: spectroscopic and enzymatic studies.
abstract: Fructose 1,6-biphosphase is a well-characterized oligomer enzyme, and many effectors allosterically control its activity. In this report, we compared the activity, allosteric properties, and conformational changes in its denaturant-induced unfolding processes. In addition, a trpytophan residue has been introduced into the interface between the C1 and C2 subunits to investigate conformational changes during unfolding. Results show that the denaturation curves of WT FruP2ase detected by various methods do not agree, and the dissociation occurs first with a monomeric form existing around 0.4 M GdmCl as shown by gel filtration. The dissociation of all mutants is accompanied by changes in fluorescence intensity. The results suggest that the unfolding of FruP2ase is a complicated, multiphase process. The activation of FruP2ase by GdmCl at low concentrations can be interpreted as a consequence of the effect of monovalent cation. In the refolding experiments, it is found that Mg2+ is not only essential for enzyme activity, but also can assist the enzyme in refolding and association by preventing the formation of aggregates.
authors.affiliation: Shanghai Institutes of Life Sciences, Chinese Academy of Sciences.
authors.forename: C
authors.initials: C
authors.lastname: Yuan
authors.forename: Z Q
authors.initials: ZQ
authors.lastname: Xie
authors.forename: F W
authors.initials: FW
authors.lastname: Zhang
authors.forename: G J
authors.initials: GJ
authors.lastname: Xu
mesh_terms: D019943:Amino Acid Substitution; D002942:Circular Dichroism; D004789:Enzyme Activation; D005453:Fluorescence; D006597:Fructose-Bisphosphatase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019791:Guanidine / Q000737:chemistry; D008274:Magnesium / Q000737:chemistry / Q000378:metabolism; D009154:Mutation; D017510:Protein Folding; D013050:Spectrometry, Fluorescence; D014364:Tryptophan / Q000737:chemistry; D014508:Urea / Q000737:chemistry
chemical_list: D014364:Tryptophan; D014508:Urea; D006597:Fructose-Bisphosphatase; D008274:Magnesium; D019791:Guanidine