Thermodynamic analysis of the structural stability of phage 434 Cro protein.
Thermodynamic parameters describing the phage 434 Cro protein have been determined by calorimetry and, independently, by far-UV circular dichroism (CD) measurements of isothermal urea denaturations and thermal denaturations at fixed urea concentrations. These equilibrium unfolding transitions are adequately described by the two-state model. The far-UV CD denaturation data yield average temperature-independent values of 0.99 +/- 0.10 kcal mol(-)(1) M(-)(1) for m and 0.98 +/- 0.05 kcal mol(-)(1) K(-)(1) for DeltaC(p)()(,U), the heat capacity change accompanying unfolding. Calorimetric data yield a temperature-independent DeltaC(p)()(,U) of 0.95 +/- 0.30 kcal mol(-)(1) K(-)(1) or a temperature-dependent value of 1.00 +/- 0.10 kcal mol(-)(1) K(-)(1) at 25 degrees C. DeltaC(p)()(,U) and m determined for 434 Cro are in accord with values predicted using known empirical correlations with structure. The free energy of unfolding is pH-dependent, and the protein is completely unfolded at pH 2.0 and 25 degrees C as judged by calorimetry or CD. The stability of 434 Cro is lower than those observed for the structurally similar N-terminal domain of the repressor of phage 434 (R1-69) or of phage lambda (lambda(6)(-)(85)), but is close to the value reported for the putative monomeric lambda Cro. Since a protein's structural stability is important in determining its intracellular stability and turnover, the stability of Cro relative to the repressor could be a key component of the regulatory circuit controlling the levels and, consequently, the functions of the two proteins in vivo.
Padmanabhan
S
S
Instituto de Estructura de la Materia, Consejo Superior de Investigaciones Cientificas, Madrid, Spain.
Laurents
D V
DV
Fernández
A M
AM
Elias-Arnanz
M
M
Ruiz-Sanz
J
J
Mateo
P L
PL
Rico
M
M
Filimonov
V V
VV
D010582:Bacteriophage lambda / Q000737:chemistry; D002152:Calorimetry, Differential Scanning; D002942:Circular Dichroism; D003090:Coliphages / Q000737:chemistry*; D004268:DNA-Binding Proteins; D006863:Hydrogen-Ion Concentration; D008956:Models, Chemical; D010446:Peptide Fragments / Q000737:chemistry; D011489:Protein Denaturation; D017510:Protein Folding; D012097:Repressor Proteins / Q000737:chemistry*; D013816:Thermodynamics; D014508:Urea / Q000737:chemistry; D014764:Viral Proteins / Q000737:chemistry*; D054334:Viral Regulatory and Accessory Proteins
D004268:DNA-Binding Proteins; D010446:Peptide Fragments; D012097:Repressor Proteins; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins; C016649:phage repressor proteins; D014508:Urea