Expression, purification, and functional characterization of recombinant human interleukin-7

Protein Expr Purif. 2009 Jan;63(1):1-4. doi: 10.1016/j.pep.2008.08.009. Epub 2008 Sep 6.

Abstract

Human interleukin-7 (IL-7) is a member of the interleukin family. Numerous studies have demonstrated IL-7's effect on B- and T-cell development as well as its potential in various clinical applications. Previously, a study reported that IL-7 could be purified from inclusion bodies using a prokaryotic system, however, the required refolding step limits the recovery rate. This study was designed to produce a bioactive recombinant human IL-7 (rhIL-7) in a eukaryotic expression system in order to obtain higher yields of the protein with simpler purification steps. We cloned human IL-7 cDNA and successfully expressed active recombinant protein in yeast using the Pichia pastoris expression system. A simple purification strategy was established to purify the rhIL-7 from the fermentation supernatant, yielding 35 mg/L at 95% purity by the use of a common SP Sepharose FF cation-exchange chromatography. Functional analysis of the purified rhIL-7 by the pre-B cell MTT (3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide) proliferation assay demonstrated a specific activity comparable to commercial sources. These results suggest that purification of rhIL-7 from yeast provides a sound strategy for large-scale production of the rhIL-7 for clinical applications as well as basic researches.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bioreactors
  • Blotting, Western
  • Chromatography, Ion Exchange
  • Cloning, Molecular
  • Fermentation
  • Humans
  • Interleukin-7 / biosynthesis*
  • Interleukin-7 / genetics
  • Interleukin-7 / isolation & purification*
  • Interleukin-7 / pharmacology
  • Pichia / genetics*
  • Pichia / metabolism
  • Protein Renaturation
  • Recombinant Proteins / biosynthesis
  • Recombinant Proteins / genetics
  • Recombinant Proteins / isolation & purification
  • Recombinant Proteins / pharmacology
  • Sepharose
  • Tetrazolium Salts
  • Thiazoles
  • Transformation, Genetic

Substances

  • IL7 protein, human
  • Interleukin-7
  • Recombinant Proteins
  • Tetrazolium Salts
  • Thiazoles
  • sepharose fast flow SP
  • Sepharose
  • thiazolyl blue