Mutational Analysis of a Red Fluorescent Protein-Based Calcium Ion Indicator
AbstractAs part of an ongoing effort to develop genetically encoded calcium ion (Ca2+) indicators we recently described a new variant, designated CH-GECO2.1, that is a genetic chimera of the red fluorescent protein (FP) mCherry, calmodulin (CaM), and a peptide that binds to Ca2+-bound CaM. In contrast to the closely related Ca2+ indicator R-GECO1, CH-GECO2.1 is characterized by a much higher affinity for Ca2+ and a sensing mechanism that does not involve direct modulation of the chromophore pKa. To probe the structural basis underlying the differences between CH-GECO2.1 and R-GECO1, and to gain a better understanding of the mechanism of CH-GECO2.1, we have constructed, purified, and characterized a large number of variants with strategic amino acid substitutions. This effort led us to identify Gln163 as the key residue involved in the conformational change that transduces the Ca2+ binding event into a change in the chromophore environment. In addition, we demonstrate that many of the substitutions that differentiate CH-GECO2.1 and R-GECO1 have little influence on both the Kd for Ca2+ and the sensing mechanism, and that the interdomain linkers and interfaces play important roles. View Full-Text
- Supplementary File 1:
Supplementary Information (PDF, 450 KB)
Scifeed alert for new publicationsNever miss any articles matching your research from any publisher
- Get alerts for new papers matching your research
- Find out the new papers from selected authors
- Updated daily for 49'000+ journals and 6000+ publishers
- Define your Scifeed now
Carlson, H.J.; Campbell, R.E. Mutational Analysis of a Red Fluorescent Protein-Based Calcium Ion Indicator. Sensors 2013, 13, 11507-11521.
Carlson HJ, Campbell RE. Mutational Analysis of a Red Fluorescent Protein-Based Calcium Ion Indicator. Sensors. 2013; 13(9):11507-11521.Chicago/Turabian Style
Carlson, Haley J.; Campbell, Robert E. 2013. "Mutational Analysis of a Red Fluorescent Protein-Based Calcium Ion Indicator." Sensors 13, no. 9: 11507-11521.