A Dynamic Interaction of Coomassie Dye with the Glycine Transporters N-termini
- 1k Downloads
Coomassie Brilliant Blue interacts with proteins and even though the interactions exhibit variation due to the amino acid content, reported dye interactions with individual proteins appear to be relatively stable. Here we report an atypical dynamic interaction of glycine transporters 1 and 2 N-termini with Coomassie dye, resulting in intramolecular interference with their Bradford assay. These proteins exhibit classic protein-Coomassie G-250 complex with absorption maximum at 595 nm, which within minutes starts to decrease and parallel increase of absorbance shoulders above 300 and 700 nm is observed. Interestingly, these effects are eliminated upon fusion of glycine transporters N-termini with glutathione S-transferase protein or by the presence of glutathione S-transferase or bovine serum albumin in the same solution. Circular dichroism data revealed largely unstructured character of glycine transporters N-termini, which suggests that dynamic properties of these protein- Coomassie complexes might be a signature of high flexibility and protein disorder. The assay might potentially reveal similar domains in other proteins and help to associate them with particular functions.
KeywordsNeurotransmitter transporters Glycine transporters Protein disorder Coomassie Brilliant Blue G-250
Glycine transporter 1
Glycine transporter 2
Bovine serum albumin
Coomassie Brilliant Blue
Sodium dodecyl sulfate
Tobacco etch virus
Polymerase chain reaction
Sodium dodecyl sulfate polyacrylamide gel electrophoresis
This work was supported by the Slovak grant agency VEGA, Grant 2/0086/13. The authors would like to thank Josef Houser PhD at the Biomolecular Interactions and Crystallization Core Facility of CEITEC for his expert technical assistance with obtaining CD spectra of proteins presented in this paper.
Compliance with Ethical Standards
Conflicts of interest
The authors declare that they have no conflicts of interest.
This article does not contain any studies with human participants or animals performed by any of the authors.
- 7.Rees MI, Harvey K, Pearce BR, Chung SK, Duguid IC, Thomas P, Beatty S, Graham GE, Armstrong L, Shiang R, Abbott KJ, Zuberi SM, Stephenson JB, Owen MJ, Tijssen MA, van den Maagdenberg AM, Smart TG, Supplisson S, Harvey RJ (2006) Mutations in the gene encoding GlyT2 (SLC6A5) define a presynaptic component of human startle disease. Nat Genet 38:801–806CrossRefGoogle Scholar
- 20.Lawrence AM, Besir HU (2009) Staining of proteins in gels with Coomassie G-250 without organic solvent and acetic acid. J Vis Exp 30:e1350Google Scholar