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KMID : 0545120110210121299
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Journal of Microbiology and Biotechnology
2011 Volume.21 No. 12 p.1299 ~ p.1305
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Combined TGE-SGE Expression of Novel PAI-1-Resistant t-PA in CHO DG44 Cells Using Orbitally Shaking Disposable Bioreactors
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Fatemeh Davami
Farzaneh Barkhordari
Mahmoud Alebouyeh
Ahmad Adeli
Fereidoun Mahboudi
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Abstract
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An important modification of thrombolytic agents is resistance to plasminogen activator inhibitor-1 (PAI-1). In previous studies, a new truncated PAI-1-resistant variant was developed based on deletion of the first three domains in t-PA and the substitution of KHRR 128-131 amino acids with AAAA in the truncated t-PA. The novel variant expressed in a static culture system of Chinese Hamster Ovary (CHO) DG44 cells exhibited a higher resistance to PAI-1 when compared with the full-length commercial drug; Actylase. In the present study, the truncatedmutant protein was expressed in CHO DG44 cells in 50 ml orbital shaking bioreactors. The final yield of the truncatedmutant in the culture was 752 IU/ml, representing a 63% increase compared with the static culture system. Therefore, these results suggest that using the combined features of a transient and stable expression system is feasible for the production of novel recombinant proteins in the quantities needed for preclinical studies.
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KEYWORD
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Tissue plasminogen activator (t-PA), Chinese Hamster Ovary (CHO), suspension culture, orbital shaker, Plasminogen activator Inhibitor-1 (PAI-1)
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