title: The conformational quality of insoluble recombinant proteins is enhanced at low growth temperatures.
abstract: Protein aggregation is a major bottleneck during the bacterial production of recombinant proteins. In general, the induction of gene expression at sub-optimal growth temperatures improves the solubility of aggregation-prone polypeptides and minimizes inclusion body (IB) formation. However, the effect of low temperatures on the quality of the recombinant protein, especially within the insoluble cell fraction, has been hardly ever explored. In this work, we have examined the conformational status of a recombinant GFP protein when produced in Escherichia coli below 37 degrees C. As expected, the fraction of aggregated protein largely decreased at lower temperatures, while the conformational quality of both soluble and aggregated GFP, as reflected by its specific fluorescence emission, progressively improved. This observation indicates that physicochemical conditions governing protein folding affect concurrently the quality of the soluble and the aggregated forms of a misfolding-prone protein, and that protein misfolding and aggregation are clearly not coincident events.
authors.affiliation: Institut de Biotecnologia i de Biomedicina, Departament de Genètica i de Microbiologia, Universitat Autònoma de Barcelona, Bellaterra, 08193 Barcelona, Spain.
authors.forename: Andrea
authors.initials: A
authors.lastname: Vera
authors.forename: Nuria
authors.initials: N
authors.lastname: González-Montalbán
authors.forename: Anna
authors.initials: A
authors.lastname: Arís
authors.forename: Antonio
authors.initials: A
authors.lastname: Villaverde
mesh_terms: D003080:Cold Temperature; D004926:Escherichia coli / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification; D002479:Inclusion Bodies / Q000737:chemistry*; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification; D017550:Spectroscopy, Fourier Transform Infrared; D013329:Structure-Activity Relationship
chemical_list: D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins