Incorporation of a tag helps to overcome expression variability in a recombinant host

Abstract

Epigenetics have witnessed a renewed interest over the past decade and assays with recombinant histones has become an important tool for uncovering various aspects of histone biology. However, at times absence of recombinant histone accumulation in bacteria is encountered which is also commonly observed for many eukaryotic proteins in general. In this study, we have investigated the effect of multiple parameters on heterologous expression of proteins. We show that there is marked variability in the accumulation of H2A.2, H2B.1, H3.2 and H4 in the recombinant host, possibly owing to translational variability and degradation by the host proteases. We found that the variability could be overcome by incorporation of the commonly used purification tags, like GST or MBP, of appropriate size and position. Our results provide compelling evidence that transcript parameters like rare codon and GC content, mRNA secondary structure etc. together modulate translation kinetics and govern recombinant protein accumulation.

Keywords

• Rare codons
• Histones
• Truncated GST
• Misfolding
• Translation