Investigation of thermal unfolding process of cyanic adduct of horseradishperoxidase by Fourier transform infrared and circular dichroism spectrometry

Detailed circular dichroism(CD) and Fourier transform infrared (FTIR) studi es have been carried out to monitor thermal unfolding of horseradish peroxi dase isoenzyme C(HRP) inhibited by CN(HRP-CN). The results suggest that HRP -CN is quite different from native HRP with different spin states of Fe of heme and different coordinated states. Cyanide becomes the sixth ligand of Fe(I) of heme and the hydrogen-binding network is destroyed partly at the s ame time, which cause the drastic decrease of thermal stability of HRP. The FTIR and Soret-CD spectra analysis demonstrate that during the heating pro cess there is an intermediate state(I') which has both partly destroyed sec ondary and tertiary structures of native HRP, then it is the appearance of protein aggregation state(A) after fully unfolding. The unfolding pathway t hus can be shown as follows: I -->I'-->U -->A.