Split-superpositive GFP reassembly is a fast, efficient, and robust method for detecting protein-protein interactions in vivo.
Split-GFP reassembly is an operationally simple in vivo technique used to identify and study interactions involving proteins and/or peptides. However, the instability of split-GFP fragments and their susceptibility to aggregation place limitations on the broader use of split-GFP reassembly. Supercharged proteins, including supercharged GFP, are variants with high theoretical negative or positive charge that are resistant to aggregation. We show that a split-superpositive GFP (split-spGFP) variant reassembles faster and more efficiently than previously reported split-sg100 GFP and split-folding-reporter GFP (split-frGFP) systems. In addition, interaction-dependent split-spGFP reassembly is efficient at physiological temperature. The increased efficiency and robustness of split-spGFP reassembly make this reporter system ideal for identifying and studying interactions involving proteins and/or peptides in vivo, and may be particularly useful for identifying or studying interactions involving proteins or peptides that are themselves susceptible to aggregation.
Department of Chemistry, Colorado State University, Fort Collins, Colorado, USA. brian.mcnaughton@colostate.edu
Brett D
BD
Blakeley
Alex M
AM
Chapman
Brian R
BR
McNaughton
D049452:Green Fluorescent Proteins / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D017433:Protein Structure, Secondary; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism*
D011506:Proteins; D049452:Green Fluorescent Proteins