Effects of succinylation on the structure and thermostability of lysozyme.
The influence of succinylation on lysozyme is studied using circular dichroism, fluorescence spectroscopy, and differential scanning calorimetry. The spectroscopic data reveal that at room temperature the structures of succinylated lysozyme and native lysozyme are similar. However, the calorimetric results show that the thermal stability of succinylated lysozyme is lower than that of native lysozyme. For succinylated lysozyme, the denaturation temperature (Td) varies in the range of 325-333 K (52-60 degrees C) and the associated denaturation enthalpy (DeltadenH) varies between 225 and 410 kJ/mol. For lysozyme, Td is 342-349 K (69-76 degrees C) and DeltadenH is 440-500 kJ/mol. From these data, the change in the heat capacity (DeltadenCp) upon thermal denaturation is derived. For lysozyme, DeltadenCp is 7.5 kJ/mol/K, and for succinylated lysozyme, it is 16.7 kJ/mol/K. The value of DeltadenCp for lysozyme is comparable to previously reported values. The high value of DeltadenCp for succinylated lysozyme is explained in terms of an extended degree of unfolding of the secondary structure and exposure of the apolar parts of the succinyl groups. Furthermore, the Gibbs energy of denaturation, as a function of temperature, derived from the thermodynamic analysis of the calorimetric data, indicates a cold-denaturated state of succinylated lysozyme below 20 degrees C. However, because a denatured state at low temperatures could not be detected by CD or fluorescence measurements, the native state may be considered to be metastable at those conditions.
Laboratory of Physical Chemistry and Colloid Science, Wageningen University, Dreijenplein 6, 6703 HB Wageningen, The Netherlands. marjin.vanderveen@wur.nl
Marijn
M
van der Veen
Willem
W
Norde
Martien Cohen
MC
Stuart
D002152:Calorimetry, Differential Scanning; D004795:Enzyme Stability; D006358:Hot Temperature; D015394:Molecular Structure; D009113:Muramidase / Q000737:chemistry*; D011489:Protein Denaturation; D013050:Spectrometry, Fluorescence; D013329:Structure-Activity Relationship; D019802:Succinic Acid / Q000737:chemistry*; D013816:Thermodynamics
D019802:Succinic Acid; D009113:Muramidase