title: Site-to-site diffusion in proteins as observed by energy transfer and frequency-domain fluorometry.
abstract: We report measurements of the site-to-site diffusion coefficients in proteins and model compounds, which were measured using time-dependent energy transfer and frequency-domain fluorometry. The possibility of measuring these diffusion coefficients were shown from simulations, which demonstrate that donor (D)-to-acceptor (A) diffusion alters the donor frequency response, and that this effect is observable in the presence of a distribution of donor-to-acceptor distances. For decay times typical of tryptophan fluorescence, the simulations indicate that D-A diffusion coefficients can be measured ranging from 10(-7) to 10(-5) cm2/s. This possibility was verified by studies of a methylene-chain linked D-A pair in solutions of varying viscosity. The D-A diffusion was also measured for two labeled peptides and two proteins, melittin and troponin I. In most cases we used global analysis of data sets obtained with varying amounts of collisional quenchers to vary the donor decay time. Unfolding of troponin I results in more rapid D-A diffusion, whereas for melittin more rapid diffusion was observed in the alpha-helical state but over a limited range of distances.
authors.affiliation: University of Maryland School of Medicine, Department of Biological Chemistry, Baltimore 21201.
authors.forename: J R
authors.initials: JR
authors.lastname: Lakowicz
authors.forename: I
authors.initials: I
authors.lastname: Gryczynski
authors.forename: J
authors.initials: J
authors.lastname: Kuśba
authors.forename: W
authors.initials: W
authors.lastname: Wiczk
authors.forename: H
authors.initials: H
authors.lastname: Szmacinski
authors.forename: M L
authors.initials: ML
authors.lastname: Johnson
mesh_terms: D004058:Diffusion; D004735:Energy Transfer; D005470:Fluorometry / Q000379:methods; D008555:Melitten / Q000737:chemistry; D008956:Models, Chemical; D011506:Proteins / Q000737:chemistry*; D014336:Troponin / Q000737:chemistry; D019210:Troponin I
chemical_list: D011506:Proteins; D014336:Troponin; D019210:Troponin I; D008555:Melitten