Effect of flexibility and cis residues in single-molecule FRET studies of polyproline.

ABSTRACT: Polyproline has recently been used as a spacer between donor and acceptor chromophores to help establish the accuracy of distances determined from single-molecule Förster resonance energy transfer (FRET) measurements. This work showed that the FRET efficiency in water is higher than expected for a rigid spacer and was attributed to the flexibility of the polypeptide. Here, we investigate this issue further, using a combination of single-molecule fluorescence intensity and lifetime measurements, NMR, theory, and molecular dynamics simulations of polyproline-20 that include the dyes and their linkers to the polypeptide. NMR shows that in water approximately 30% of the molecules contain internal cis prolines, whereas none are detectable in trifluoroethanol. Simulations suggest that the all-tr

SUBMITTER: Best RB 

PROVIDER: S-EPMC2141891 | biostudies-literature | 2007 Nov

REPOSITORIES: biostudies-literature