The conformational quality of insoluble recombinant proteins is enhanced at low growth temperatures

• Published in: Biotechnology and bioengineering Vol. 96; no. 6; pp. 1101 - 1106
• Main Authors: Vera, Andrea, González-Montalbán, Nuria, Arís, Anna, Villaverde, Antonio
• Format: Journal Article
• Language: English
• Published: Hoboken Wiley Subscription Services, Inc., A Wiley Company 15-04-2007; Wiley; Wiley Subscription Services, Inc
• Subjects: ATR; Biological and medical sciences; Biotechnology; Cold Temperature; E coli; Escherichia coli; Escherichia coli - metabolism; Fluorescence; Fundamental and applied biological sciences. Psychology; Gene expression; GFP; GFP, ATR; Green Fluorescent Proteins - biosynthesis; Green Fluorescent Proteins - chemistry; Green Fluorescent Proteins - isolation & purification; inclusion bodies; Inclusion Bodies - chemistry; inclusion bodies, protein aggregation; Peptides; protein aggregation; Protein Conformation; protein quality; Proteins; Recombinant Proteins - biosynthesis; Recombinant Proteins - chemistry; Recombinant Proteins - isolation & purification; Spectroscopy, Fourier Transform Infrared; Structure-Activity Relationship; Temperature effects; Aggregation; Growth; Quality; Green fluorescent protein; inclusion bodies; Inclusion body; Recombinant protein; Low temperature; Conformation; ATR; protein aggregation; E. coli; protein quality; GFP
• ISSN: 0006-3592; 1097-0290
• DOI: 10.1002/bit.21218

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°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. Biotechnol. Bioeng. 2007;96:1101–1106. © 2006 Wiley Periodicals, Inc.