Part:BBa_K2926048

mCherry with hexahistidine tag for purification

This part codes for the red fluorescent protein mCherry with a C-terminal hexahistidine tag for simple purification via metal ions.

Usage and Biology

The red fluorescent protein mCherry was fused to a hexahistidine tag to enable easy purification. Since the first successful cloning of the green fluorescent protein GFP of Aequorea victoria in 1992 (Prasher et al. 1992) fluorescent proteins became a widely used tool in many fields of research. In contrast to antibodies labeled with fluorophores that have to cross the cellular membrane which severely disturbes the cellular integrity, flourescing proteins enable live cell imaging and the investigation of native states of the cell.
Because of the wide range of applications for fluorescing proteins there was a great interest in finding and engineering improved variants and a wider colour spectrum. In the last few years red fluorescing proteins became more and more important. Common native red fluorescing proteins are often dimeric or tetrameric what makes their usage in experimental setups difficult. Directed mutation of dsRFP from the corallimorpharia Discosoma sp. Led to the first monomeric red fluorescing protein mRFP1 (Shaner et al. 2004). Unfortunately this mutations resulted in a lower quantum yield and decreased photostability (Shaner et al. 2004). During further protein engineering attempts, scientists were able to create the red fluorescent protein mCherry. mCherry is a 26.7  kDa protein that shows a very short maturation time of about 15  minutes and a low acid sensitivity. Its excitation maximum lies at 587  nm and it has its emission maxiumum at 610  nm (www.fpbase.org). In 2006 the crystal structure of mCherry was published (Shu and Remington 2006).

Crystal structure of mCherry (Shu and Remington 2006).

mCherry consists of 13 beta-sheets which form a beta-barrel and three alpha helices. The chromophore is made of methionine, tyrosine and glycine who posttranslationally form an imidazolinone (Shu et al. 2006).