The stability of enzymes remains a critical issue in biotechnology. Compared with the strategies for obtaining stable enzymes, chemical modification is a simple and effective technique. In the present study, chemical modification of horseradish peroxidase (HRP) was carried out with pyromellitic anhydride. HRP has achieved a prominent position in the pharmaceutical, chemical, and biotechnological industries. In this study, the effect of chemical modification on thermal stability, structure, and function of the enzyme was studied by fluorescence, circular dichroism, and absorbance measurements. The results indicated a decrease in compactness of the structure and a considerable enhancement in thermal stability of HRP below 60 A degrees C. It seems the charge replacement and introduction of the bulky group bring about the observed structural and the functional changes.