Calcium triggers the refolding of Bacillus subtilis chitosanase.
We characterized the reversible folding-unfolding transition of Bacillus subtilis exocellular chitosanase from either thermal or urea denaturation of the protein. The transitions were monitored in each case by intrinsic fluorescence changes and resistance to proteolysis. Unfolding and refolding kinetics and differential scanning calorimetry analysis suggested a two-state equilibrium. The equilibrium between the folded and unfolded states was rapidly displaced towards the folded state in the presence of a low concentration of calcium (2-20 mM). The binding titration curve indicated that chitosanase possesses one weak Ca(2+)-binding site (with an equilibrium affinity constant, K (A), of 0.3x10(3) M(-1)). These results support the hypothesis that this metal ion, which is accumulated in the cell wall environment of B. subtilis, is an effector that influences folding and stability of newly translocated proteins.
Colomer-Pallas
Anne
A
Institut Jacques Monod, Laboratoire Génétique et Membranes, Centre National de la Recherche Scientifique, Universités Paris 6 et Paris 7, Tour 43, 2 place Jussieu, 75251 Paris cedex 05, France.
Pereira
Yannick
Y
Petit-Glatron
Marie-Françoise
MF
Chambert
Régis
R
D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000201:enzymology* / Q000254:growth & development; D001426:Bacterial Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D001665:Binding Sites; D002118:Calcium / Q000737:chemistry / Q000378:metabolism*; D002455:Cell Division / Q000502:physiology; D017886:Durapatite / Q000378:metabolism; D004795:Enzyme Stability; D005453:Fluorescence; D006026:Glycoside Hydrolases / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D017186:Titrimetry; D014508:Urea / Q000737:chemistry
D001426:Bacterial Proteins; D014508:Urea; D017886:Durapatite; D006026:Glycoside Hydrolases; C019868:chitosanase; D002118:Calcium