PEGylation of deuterohaemin–alanine–histidine–threonine–valine–glutamic acid–lysine and its influence on activity, stability, and aggregation

Deuterohaemin–alanine–histidine–threonine–valine–glutamic acid–lysine (DhHP-6) is a synthetic heme-containing peroxidase mimic that exhibits a high peroxidase enzyme activity. Compared to other microperoxidases, DhHP-6 has a poor stability and tends to aggregate in aqueous solutions. In this study, poly(ethylene glycol) (PEG) was used to improve the properties of DhHP-6. Factors that affected the PEGylation product yield were investigated. PEGylated DhHP-6 (mPEG–DhHP-6) was characterized by reversed-phase high-pressure liquid chromatography (RP-HPLC), matrix-assisted laser desorption/ionization time of flight mass spectra (MALDI-TOF-MS), and ultraviolet–visible (UV–vis) spectroscopy. The results show that the optimal PEGylation reaction conditions were achieved when the PEGylation was conducted in a borate buffer solution at pH 8.0 and 25°C for 4 h with a feeding ratio of 2 equiv of active PEG. After PEGylation, mPEG–DhHP-6 showed a great improvement in its stability with little activity loss. The UV–vis spectra of DhHP-6 and mPEG–DhHP-6 in different pH solutions showed that the aggregation of DhHP-6 was partly suppressed after PEGylation. © 2012 Wiley Periodicals, Inc. J. Appl. Polym. Sci., 2013