title: Production of soluble eukaryotic recombinant proteins in E. coli is favoured in early log-phase cultures induced at low temperature.
abstract: BACKGROUND
  Producing recombinant plant proteins expressed in Escherichia coli produce in high yields and in a soluble and functional form can be difficult. Under overexpression conditions, proteins frequently accumulate as insoluble aggregates (inclusion bodies) within the producing bacteria. We evaluated how the initial culture density, temperature and duration of the expression stage affect the production of some eukaryotic enzymes in E. coli.
  
  
  RESULTS
  A high yield of active soluble proteins was obtained by combining early-log phase cultures and low temperatures for protein induction. When IPTG was added at OD600 = 0.1 and cultures were maintained at 4°C for 48-72 h, the soluble protein yield was 3 fold higher than that obtained in the mid-log phase (OD600 = 0.6). Besides, the target protein expression increased and the endogenous bacterial proteins reduced, thus making the protein purification process easier and more efficient.
  
  
  CONCLUSIONS
  The protocol can be widely applied to proteins with a heterologous expression which was limited by loss of activity at high temperatures or by low soluble recombinant protein yield.
authors.affiliation: Departamento de Biología Vegetal, Facultad de Farmacia, Universidad de Valencia, Av. VA Estellés s/n, Burjasot, 46100 Spain.
authors.forename: Teresa
authors.initials: T
authors.lastname: San-Miguel
authors.forename: Pedro
authors.initials: P
authors.lastname: Pérez-Bermúdez
authors.forename: Isabel
authors.initials: I
authors.lastname: Gavidia
keywords: Early log phase; Functional proteins; Low temperature; Soluble recombinant proteins