Response of rubredoxin from Pyrococcus furiosus to environmental changes: implications for the origin of hyperthermostability.
The bases of the hyperthermostability of rubredoxin from Pyrococcus furiosus (RdPf) have been probed by structural perturbations induced by solution pH and ionic strength changes. Comparison of the solution behavior at pH 7 and pH 2, as probed by far- and near-UV circular dichroism, Trp fluorescence emission, 1-anilinonaphthalene-8-sulfonate (ANS) binding, and NMR spectroscopy, reveals the presence of only minimal structural variations at room temperature. At pH 2, the protein displays a surprising nearly native-like behavior at high ionic strength while, at low ionic strength, it is capable of strongly binding the hydrophobic probe ANS. All the secondary and tertiary structural features, including the environment of the hydrophobic core, appear to be intact regardless of pH and ionic strength. The apparent "melting" or denaturation temperature at pH 2, however, is 42 degrees C lower than at pH 7. This is attributed to the perturbation of many electrostatic interactions, including the disruption of all the ion pairs, which is complete at pH 2, as indicated by electrometric pH titrations. The implications of these findings for the origins of the hyperthermostability of rubredoxin are discussed.
Cavagnero
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Arthur Amos Noyes Laboratories of Chemical Physics, California Institute of Technology Pasadena, 91125, USA.
Zhou
Z H
ZH
Adams
M W
MW
Chan
S I
SI
D000817:Anilino Naphthalenesulfonates / Q000378:metabolism; D001105:Archaea / Q000737:chemistry*; D002942:Circular Dichroism; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D008958:Models, Molecular; D009994:Osmolar Concentration; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012416:Rubredoxins / Q000737:chemistry* / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013053:Spectrophotometry; D013499:Surface Properties; D017186:Titrimetry
D000817:Anilino Naphthalenesulfonates; D012416:Rubredoxins; C027132:1-anilino-8-naphthalenesulfonate