{"id":21927,"date":"2020-03-16T10:52:55","date_gmt":"2020-03-16T09:52:55","guid":{"rendered":"https:\/\/www.neb-online.de\/?page_id=21927"},"modified":"2023-12-19T14:21:06","modified_gmt":"2023-12-19T13:21:06","slug":"nebexpress-mbp-fusion-and-purification-system","status":"publish","type":"page","link":"https:\/\/www.neb-online.de\/en\/proteinexpression\/nebexpress-mbp-fusion-and-purification-system\/","title":{"rendered":"NEBExpress MBP Fusion and Purification System"},"content":{"rendered":"

[vc_row][vc_column][vc_basic_grid post_type=”post” max_items=”10″ element_width=”12″ gap=”15″ item=”4123″ taxonomies=”68″ grid_id=”vc_gid:1702991912476-165ae54c-04ca-5″][\/vc_column][\/vc_row][vc_row bwd_show_when_date=”03\/11\/2020 23:00″ bwd_tbc4vd_show_weekday=””][vc_column width=”1\/2″][vc_column_text]The NEBExpress MBP Fusion and Purification System takes advantage of the strong Ptac promoter and the translation initiation signals of maltose binding protein (MBP) to enhance solubility and expression levels of a desired protein in E. coli<\/em>.<\/p>\n

The resulting product is an MBP fusion protein, which is then purified by affinity chromatography.<\/p>\n

The included pMAL-c6T vector expresses the N-terminal hexahistidine tagged malE gene (lacking its secretory signal sequence and engineered for tighter binding to amylose) followed by a multiple cloning site containing a TEV protease recognition sequence and stop codons in all three frames. The pMAL-c6T vector expresses the MBP fusion in the cytoplasm.[\/vc_column_text][\/vc_column][vc_column width=”1\/2″][vc_text_separator title=”NEBExpress MBP System – Workflow” color=”custom” accent_color=”#eb7e2d”][vc_single_image image=”21655″ img_size=”full”][\/vc_column][\/vc_row][vc_row bwd_show_when_type=”after” bwd_show_when_date=”03\/11\/2020 23:00″ bwd_tbc4vd_show_weekday=”” disable_element=”yes”][vc_column width=”1\/2″][vc_column_text]Das neue NEBExpress MBP Fusion and Purification System nutzt den f\u00fcr Proteinexpression in E. coli<\/em> optimierten Ptac Promoter in Kombination mit der Sequenz des Maltose-Binding-Protein (MBP).<\/p>\n

Das resultierende MBP-Fusionsprotein kann in gro\u00dfen Mengen im Cytoplasma von E. coli<\/em> exprimiert werden. Das MBP-Tag dient dabei zum einen der einfachen Aufreinigung \u00fcber ein Amylose S\u00e4ule, erh\u00f6ht aber auch die L\u00f6slichkeit des Proteins in der Zelle. So entstehen deutlich weniger Probleme mit unlo\u0308slichen Produkten, wie z.B. “inclusion bodies”.[\/vc_column_text][\/vc_column][vc_column width=”1\/2″][vc_text_separator title=”NEBExpress MBP System – Workflow” color=”custom” accent_color=”#eb7e2d”][vc_single_image image=”21834″ img_size=”full”][\/vc_column][\/vc_row][vc_row][vc_column][vc_column_text]The fusion protein can be purified by a one-step purification method using an amylose matrix and MBPs affinity for maltose. Small-scale protein purification can directly be done from 200 – 500 \u00b5l culture supernatant, while large-scale purifications are possible from up to 1000 ml of bacterial culture.[\/vc_column_text][\/vc_column][\/vc_row][vc_row][vc_column width=”1\/2″][vc_column_text]Following amylose purification, the target protein can be cleaved from the MBP-tag using TEV Protease, without adding any vector-derived residues to the protein. Both the MBP-tag and TEV Protease are polyhistidine-tagged for easy removal from the reaction. The target protein yield can be up to 100 mg\/l, with typical yields in the range of 10\u201340 mg\/l.[\/vc_column_text][\/vc_column][vc_column width=”1\/2″ css=”.vc_custom_1584352165533{margin-bottom: 20px !important;}”]

\n\t\t\t\tAdvantages:<\/span>\n\t\t\t\t\t