Liming Ying

Peptide synthesis PAK1 activating peptide (PAP) TSNSQKYMSFTDKSA was prepared as previously described.28 Briefly, PAP derived from the PAK1 autoinhibitory region was linked to the 11-amino acid sequence YGRKKRRQRRR derived from HIV-1 trans-activating regulatory protein. The peptide (YGRKKRRQRRRGTSNSQKYMSFTDKSA) was synthesized in the proteomics core lab in Research Resource Centre at University of Illinois at Chicago (UIC), USA and was confirmed by mass spectrometry.

Tetramethylrhodamine self-quenching is a probe of conformational change on the scale of 15 – 25 Å Tetramethylrhodamine (TMR) is a fluorescent dye whose self-quenching has been used as a probe of multiple biological phenomena. We determine the distance-dependence of self-quenching and place bounds on the timescale of TMR dissociation. Our results validate fluorescence self-quenching as an alternative to FRET and enable future assays to be designed with confidence. This article is Open Access

Abstract Tetramethylrhodamine (TMR) is a fluorescent dye whose self-quenching has been used as a probe of multiple biological phenomena. We determine the distance-dependence of self-quenching and place bounds on the timescale of TMR dissociation. Our results validate fluorescence self-quenching as an alternative to FRET and enable future assays to be designed with confidence. Competing Interest Statement The authors have declared no competing interest.