Engineering of chimeric catalase-Angiopep-2 for intracellular protection of brain endothelial cells against oxidative stress

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• 作者：Sakda Yainoy^a ; ^b ; Patcharaporn Houbloyfa^b ; Warawan Eiamphungporn^a ; ^b ; Chartchalerm Isarankura-Na-Ayudhya^a ; Virapong Prachayasittikul^a ; ^{virapong.pra@mahidol.ac.th" class="auth_mail}
• 关键词：Catalase ; Angiopep-2 ; Blood&ndash ; brain barrier ; Brain microvascular endothelial cells ; Oxidative stress ; Fusion protein
• 刊名：International Journal of Biological Macromolecules
• 出版年：July 2014
• 年：2014
• 卷：68
• 期：Complete
• 页码：60-66
• 全文大小：1215 K

文摘

Blood–brain barrier (BBB) disruption and brain microvascular endothelial cells (BMVECs) death caused by excessive production of hydrogen peroxide (H₂O₂) have been implicated in several neurological conditions. To overcome this problem, H₂O₂-degrading enzyme with ability to enter the BMVECs is required. In the present study, genetic fusion of gene encoding human catalase and gene encoding Angiopep-2 (AP2), a brain targeting peptide, was performed. The fusion protein was successfully expressed in Escherichia coli and purified to homogeneity. The protein retained heme content and specific enzymatic activity in the same order of magnitude as that of native enzyme. Study of the BMVECs internalization showed that 0.1 渭M of the fusion protein can enter the cell within 15 min, while internalization of the native protein was not observed at this condition. In addition, treatment of the BMVECs with 20 units of the fusion protein for 30 min showed protection against H₂O₂ up to 5.0 mM, whereas this protective effect was not observed from treatment with the native protein. Therefore, construction of chimeric human catalase and AP2 provides an insight into the development of potential therapeutic antioxidant with ability to penetrate the BBB for protection against neurodegenerative disorders.