Overexpression and purification of single zinc finger peptides of human zinc finger protein ZNF191

Dong Xin Zhao, Zhi Chun Ding, Yu Qi Liu, Zhong Xian Huang

Research output: Contribution to journalArticle

10 Scopus citations

Abstract

ZNF191, a new human zinc finger protein, probably relates to some hereditary diseases and cancers. To obtain structural information of zinc finger domain a convenient method for obtaining milligram quantities of each zinc finger peptide of ZNF191 is necessary. Here, we report an Escherichia coli expression system for rapid and high-level expression of zinc finger 3 and zinc finger 4 of ZNF191. The gene of zinc finger 3 or zinc finger 4 was cloned into pET31b vector to allow expression of single zinc finger peptide as a ketosteroid isomerase (KSI) fusion protein. The KSI-single zinc finger fusion protein was overexpressed in the form of inclusion body, which can be purified by washing several times using buffer solutions, and then be cleaved directly by cyanogen bromide to release single zinc finger peptide. The more than 20 mg/L yield of single zinc finger peptide was achieved with more than 95% purity by using YM ultrafiltration membranes. Circular dichroism spectra of these two single zinc finger peptides titrated with Zn2+ ions demonstrate that they have different secondary structures.

Original languageEnglish (US)
Pages (from-to)232-237
Number of pages6
JournalProtein Expression and Purification
Volume53
Issue number1
DOIs
StatePublished - May 1 2007
Externally publishedYes

Keywords

  • Cyanogen bromide cleavage
  • Ketosteroid isomerase
  • Zinc finger protein

ASJC Scopus subject areas

  • Biotechnology

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