当前位置： SCI文献检索 > INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES期刊下所有文献 > Enhancing chimeric hydrophobin II-vascular endothelial growth factor A165 expression in Pichia pastoris and its efficient purification using hydrophobin counterpart.

Enhancing chimeric hydrophobin II-vascular endothelial growth factor A165 expression in Pichia pastoris and its efficient purification using hydrophobin counterpart.

Abstract：

:We report cloning and expressing of recombinant human VEGF-A165, fused at the N-terminal with Hydrophobin II (HFBII) from Trichoderma reseei, in yeast Pichia pastoris. We validated the construct using SDS-PAGE and ELISA against VEGF-A165 and efficiently performed protein purification and enrichment based on HFBII counterpart and using an aqueous two-phase system (ATPS) with nonionic surfactant X-114. We studied the effects of various culture medium additives and interaction effects of positive factors to increase the recombinant HFBII-VEGF-A165 production. Supplementing the Pichia pastoris cell culture medium with Mg2+, Polysorbate 20 (PS 20), and 4-phenylbutyrate (PBA) improved the expression of the chimeric protein. Orthogonal experiments showed that the optimal condition to achieve maximal HFBII-VEGF-A165 production was with the addition of PBA, PS 20, and MgSO4. Under this condition, the production of the target protein was 4.5 times more than that in the medium without the additives. Overall, our approach to produce chimeric HFBII-VEGF-A165 and selectively capture it in ATPS is promising for large-scale protein production without laborious downstream processing.

journal_name

Int J Biol Macromol

journal_title

International journal of biological macromolecules

authors

Arjmand S,Tavasoli Z,Ranaei Siadat SO,Saeidi B,Tavana H

doi

10.1016/j.ijbiomac.2019.08.080

subject

Has Abstract

pub_date

2019-10-15 00:00:00

pages

1028-1034

eissn

0141-8130

issn

1879-0003

pii

S0141-8130(19)33563-9

journal_volume

139

pub_type

杂志文章

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