Excerpts

StayGold is an exceptionally bright and stable fluorescent protein that is highly resistant to photobleaching. Despite favourable fluorescence properties, use of StayGold as a fluorescent tag is limited because it forms a natural dimer. Here we describe the 1.6 Å structure of StayGold and generate a new derivative (mStayGold) that retains the brightness and photostability of the original protein while being fully monomeric.

to establish monomerization in vivo, we performed the organized smooth endoplasmic reticulum (OSER) assay for StayGold and mStayGold E138D alongside dTomato (an established dimer) and mTurquoise (a known monomer; Supplementary Fig. 14). The data show that mStayGold E138D is monomeric, with an OSER score comparable to that of the mTurquoise control, while the original StayGold is a weak dimer, with an OSER score that is lower than mStayGold but higher than dTomato. The apparent fragility of the StayGold dimer in the OSER assay is consistent with our observation that the interface can easily be broken with single amino acid substitutions.