Browse views: by Year, by Function, by GLF, by Subfunction, by Conference, by Journal

High-level Soluble Expression, Purification and Characterization of Active Human Midkine from Escherichia coli

Yan, Wei and Goette, Marjo and Hofmann, Gabriele and Zaror, Isabel and Sim, Bang Janet (2010) High-level Soluble Expression, Purification and Characterization of Active Human Midkine from Escherichia coli. Protein Expression and Purification, 70 (2). pp. 270-276. ISSN 1046-5928

Abstract

Midkine (MDK) belongs to a class of heparin-binding growth factors and is highly expressed in a number of cancers. MDK is a cysteine-rich 13 kDa protein containing five disulfide bonds. In this study, we expressed recombinant human MDK (rhMDK) in Escherichia coli Origami 2 (DE3) strain, which carries a (trxB-/gor522-) double mutation. Soluble rhMDK was expressed at a high level in this strain and the protein was purified by a two-step purification using heparin affinity and gel filtration chromatography. Seven milligrams of rhMDK with high purity were obtained from a 3 L culture. All 10 cysteines were confirmed to be engaged in correct disulfide bond linkages by mass spectrometry analysis. Activity of purified rhMDK was confirmed by a neurite outgrowth assay using rat cerebellar granule cells. Active rhMDK is a critical reagent for cancer drug discovery studies.

Item Type: Article
Related URLs:
Additional Information: author can archive post-print (ie final draft post-refereeing); Publisher's version/PDF cannot be used
Keywords: Human midkine; Disulfide bond; Escherichia coli expression; Affinity purification; Analysis
Related URLs:
Date Deposited: 13 Oct 2015 13:16
Last Modified: 13 Oct 2015 13:16
URI: https://oak.novartis.com/id/eprint/1926

Search

Email Alerts

Register with OAK to receive email alerts for saved searches.