Distance distributions in native and random-coil troponin I from frequency-domain measurements of fluorescence energy transfer

✍ Scribed by Joseph R. Lakowicz; Ignacy Gryczynski; Herbert C. Cheung; Chien-Kao Wang; Michael L. Johnson

• Publisher: Wiley (John Wiley & Sons)
• Year: 1988
• Tongue: English
• Weight: 523 KB
• Volume: 27
• Category: Article
• ISSN: 0006-3525
• DOI: 10.1002/bip.360270509

✦ Synopsis

We used time-dependent fluorescence energy transfer to determine the distribution of donorto-acceptor distances in native and denatured troponin I(Tn1). The single tryptophan residue (Trp 158) of TnI served as the donor (D), and the acceptor (A) was a labeled cysteine residue (Cys 133). The time-dependent intensity decays of the donor were measured by the frequency-domain method from 10 to 320 MHz. T h e frequency response of the donor emission, in the absence and presence of acceptor, was used to recover the distribution of D to A distances, using an algorithm that accounts for the intrinsic multiexponential decay of tbe donor. In the native st@e the D-A distribution is characterized by an average distance of 23 A and aQhalf-width of 12 A. Denaturation results in a modest increase in the average distance to 27 A, and a dramatic increase in half-width to 47 A. We believe the ability to recover distance distributions will have numerous applications in the characterization of biological macromolecules.