Περίληψη:
We have studied the stability of the histone-like, DNA-binding protein
HU from the hyperthermophilic eubacterium Thermotoga maritima and its
E34D mutant by differential scanning microcalorimetry and CD under
acidic conditions at various concentrations within the range of 2-225
mum of monomer. The thermal unfolding of both proteins is highly
reversible and clearly follows a two-state dissociation/unfolding model
from the folded, dimeric state to the unfolded, monomeric one. The
unfolding enthalpy is very low even when taking into account that the
two disordered DNA-binding arms probably do not contribute to the
cooperative unfolding, whereas the quite small value for the unfolding
heat capacity change (3.7 kJ.K-1.mol(-1)) stabilizes the protein within
a broad temperature range, as shown by the stability curves (Gibbs
energy functions vs. temperature), even though the Gibbs energy of
unfolding is not very high either. The protein is stable at pH 4.00 and
3.75, but becomes considerably less so at pH 3.50 and below, to the
point that a simple decrease in concentration will lead to unfolding of
both the wild-type and the mutant protein at pH 3.50 and low
temperatures. This indicates that various acid residues lose their
charges leaving uncompensated positively charged clusters. The wild-type
protein is more stable than its E34D mutant, particularly at pH 4.00 and
3.75 although less so at 3.50 (1.8, 1.6 and 0.6 kJ.mol(-1) at 25
degreesC for DeltaDeltaG at pH 4.00, 3.75 and 3.50, respectively), which
seems to be related to the effect of a salt bridge between E34 and K13.
Συγγραφείς:
Ruiz-Sanz, J
Filimonov, VV
Christodoulou, E
Vorgias, CE and
Mateo, PL
