A novel approach to the expression and purification of recombinant Xenopus Cdc25C

Stephanie Ramadan, Ghizlaine Bendriss, Satanay Zuhair Hubrack, Abdulrahman Al-Abdulmalek, Khaled Machaca

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

The Cdc25 family encodes dual specificity protein phosphatases that play critical roles in cell cycle progression. Activation of the Cdc25C represents a primary driver for meiosis progression in Xenopus oocytes. Given its central role in meiosis the Xenopus Cdc25C has been studied extensively, however purification of the recombinant protein is difficult thus preventing better characterization of its function. Here we describe methods to overcome these difficulties resulting in the production of high purity and yield recombinant Xenopus Cdc25C. We use a synthetic Xenopus Cdc25C gene that was codon optimized for expression in E. coli. We further combine an N-terminal His-tag with a C-terminal Strep-tag II, to isolate extremely pure full-length Cdc25C protein. The recombinant Xenopus Cdc25C is active both in vitro using a phosphatase assay and in vivo when injected into Xenopus oocytes. This new approach should be applicable to the purification of other members of the Cdc25 gene family.

Original languageEnglish
Pages (from-to)148-152
Number of pages5
JournalProtein Expression and Purification
Volume120
DOIs
Publication statusPublished - 1 Apr 2016

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Keywords

  • Cdc25C
  • Cell cycle
  • E. coli optimized
  • Oocyte
  • Phosphatase
  • Xenopus

ASJC Scopus subject areas

  • Biotechnology

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