Examining the Polyproline Nanoscopic Ruler in the Context of Quantum Dots

Abstract

The rigidity and defined length of the polyproline type II helix (PPII) have made it the structural basis of a nanoscopic ruler, which has been widely applied in Forster resonance energy transfer (FRET) studies. A growing body of data, however, has questioned the foundation for this and has provided evidence for structural perturbations to the PPII caused by temperature, salt content, solvent polarity, and even Pro repeat length. Here, we examine the polyproline ruler in the context of semiconductor quantum dots (QDs) and FRET. For this study, a series of polyproline peptides (Pron, n = 0, 3, 6, 9, 12, 15, 18) displaying a C-terminal hexahistidine sequence (His6) and an N-terminal cysteine for site-specific labeling with Cy3 dye were synthesized. Peptide rigidity was first examined with ATTO 647 Ni2+-nitrilotriacetic acid acceptor dye coordinated to the His6-termini of the Cy3 donor-labeled peptides. These conditions provided a steady-state fluorescent response that closely followed FRET predictions deriv...