SECRETION OF BACILLUS-SUBTILIS LEVANSUCRASE - FE(III) COULD ACT AS A COFACTOR IN AN EFFICIENT COUPLING OF THE FOLDING AND TRANSLOCATION PROCESSES

被引:22
作者
CHAMBERT, R
BENYAHIA, F
PETITGLATRON, MF
机构
[1] Institut Jacques Monod, Lab. Genetique et Membranes, Universite Paris 7, 75251 Paris Cedex 05
关键词
D O I
10.1042/bj2650375
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The refolding of levansucrase denatured by urea was studied as a possible model for the second step of the secretion pathway of this protein. The folding-unfolding transition was monitored by measuring intrinsic fluorescence and resistance to proteolysis. Both methods provided the same estimation for the unfolding free energy of levansucrase, ΔG(D), which was 30.1 ± 1.7 kJ · mol-1) (7.2 ± 0.4 kcal · mol-1) at pH 7 in 0.1 M-potassium phosphate buffer. The rate of refolding was greatly enhanced by Fe3+, whereas the Fe3+ chelator EDTA prevented correct refolding. Fe3+ allowed the protein to reach its folded form in medium in which the dielectric constant had been lowered by ethanol. The efficiency in vivo of the export of levansucrase bearing an amino acid modification which blocks the second step of the translocation pathway was greatly increased by high concentrations of Fe3+ in the culture medium. Assuming that the protein folding governs the second step of the secretion process of levansucrase, we discuss from an irreversible thermodynamic point of view the possible role of Fe3+ in the efficient coupling of the two events.
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页码:375 / 382
页数:8
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