Characterization and stability investigation of Cu,Zn-superoxide dismutase covalently modified by low molecular weight heparin

Abstract

Cu,Zn-superoxide dismutase (SOD) was chemically modified with low molecular weight heparin (LMWH). To characterize the conjugate, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and native polyacrylamide gel electrophoresis (native PAGE) with protein staining and polysaccharide staining were employed. The stabilities of the modified enzyme to heat, acid, alkali, and trypsin treatment were also investigated. SDS-PAGE of the conjugate presented two major bands, and native PAGE of the conjugate showed similar banding position with protein staining and polysaccharide staining, which was different from that of the unmodified SOD and LMWH/SOD mixture. Moreover, the conjugate migrated faster with increasing extent of the modification. Enhanced heat stability, acid resistance, alkali resistance, and anti-trypsin stability of the modified enzyme were observed compared with those of the unmodified enzyme. Results of the study suggest that covalent linkage in LMWH-SOD can be effectively characterized by electrophoretic techniques and the chemical modification of SOD with LMWH can enhance the stabilities of the enzyme. In addition, native PAGE with protein staining can be used to evaluate the extent of the modification.

Research progress in the radioprotective effect of superoxide dismutase

“…SOD conjugates obtained through the way of chemical modifi cation have advantages of prolonged halflife, improved cell membrane permeability, augmented bioactivities and effi cient targeting compared with the native SOD. An outstanding example of these compounds is Hep-SOD which has been verifi ed for its superior radioprotection owing to its long half-life, and enhanced tolerance to high temperature, strong acid/base, and enzymolysis of trypsin (67,68). Besides these advanced forms of SOD, nonenzymatic SOD mimics have also become a favorite form of radioprotective agent to some researchers in recent years.…”

Approximation of new generation pharmacological enzyme researches to clinical practice

“…В зависимости от соотношения весовых концентраций альбумина и полимерного носителя образуется два типа конъюгатов: белок в плотном полимерном окружении в структуре частицы конъюгата или частицы конъюгата, имеющие более рыхлую структуру, в которой белок практически экспонирован в растворитель. Ковалентное присоединение низкомолекулярного гепарина к Cu, Znсупероксиддисмутазе способствовало получению ферментного термостабильного конъюгата с резистентностью к действию низких и высоких значений рН среды и устойчивостью к трипсиновому протеолизу [18]. Для преодоления резистентности к действию рекомбинантной аргининдеиминазы при аргинин-ауксотрофическом раке использо-А.В.…”

The radiation protection role of heparin-SOD conjugate in irradiated mice

“…Hep-SOD conjugate was synthesized according to the method reported previously Zhang et al, 2006). The protein content and enzymatic activity of Hep-SOD were determined by BCA assay kit and SOD assay kit, respectively.…”