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J. Microbiol. Biotechnol. 2010; 20(10): 1430-1435

Published online October 28, 2010 https://doi.org/10.4014/jmb.1004.04040

Copyright © The Korean Society for Microbiology and Biotechnology.

The Orientation-dependent expression of angiostatin-endostatin hybrid proteins and their characterization for the synergistic effects of antiangiogenesis

Sun-Yeol Paek 1, Yong-Seok Kim 2 and Shin-Geon Choi 1*

1Department of Bioengineering and Technology, Kangwon National University, Chuncheon 200-701, Korea, 2Department of Biochemistry and Molecular Biology, College of Medicine, Hanyang University, Seoul 133-791, Korea

Received: April 30, 2010; Accepted: July 5, 2010

Abstract

Two angiostatic fusion proteins (hAE and hEA) of human angiostatin (hAS) and endostatin (hES) proteins differed in tandem connection manner were constructed and evaluated for synergistic anti-angiogenic effects. The 65 kDa secreted fusion proteins from Pichia pastoris expression were verified by mass-spec analysis and western blotting assay. Luciferase reporter gene assay using VEGF promoter revealed that angiostatin-endostatin fusion protein (hAE) and its corresponding fusion gene delivery on Human Microvascular Endothelial Cells (HMEC 1) resulted in more potent synergistic anti-angiogenic effects than endostatin-angiostatin fusion protein (hEA). These facts suggest that the orientation of fusion genes between hAS and hES might be an important factor for developing therapeutic proteins.

Keywords: angiogenesis inhibitor, angiostatin, endostatin, fusion protein, Pichia expression