Association and activation of fructose 1,6-bisphosphase during unfolding and refolding: Spectroscopic and enzymatic studies

Citation
C. Yuan et al., Association and activation of fructose 1,6-bisphosphase during unfolding and refolding: Spectroscopic and enzymatic studies, J PROTEIN C, 20(1), 2001, pp. 39-47
Citations number
29
Categorie Soggetti
Biochemistry & Biophysics
Journal title
JOURNAL OF PROTEIN CHEMISTRY
ISSN journal
02778033 → ACNP
Volume
20
Issue
1
Year of publication
2001
Pages
39 - 47
Database
ISI
SICI code
0277-8033(200101)20:1<39:AAAOF1>2.0.ZU;2-R
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 dena turant-induced unfolding processes. In addition, a trpytophan residue has b een introduced into the interface between the C1 and C2 subunits to investi gate conformational changes during unfolding. Results show that the denatur ation curves of WT FruP(2)ase 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 accomp anied by changes in fluorescence intensity. The results suggest that the un folding of FruP(2)ase is a complicated, multiphase process. The activation of FruP(2)ase by GdmCl at low concentrations can be interpreted as a conseq uence 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.