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Proceedings Paper

Characterization of triplet probes on filamentous actin
Author(s): Choi-man Ng; Richard D. Ludescher
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Paper Abstract

We have used steady-state measurements of polarized emission to determine the orientation of a xanthene probe covalently attached to cys374 in F-actin oriented by flow. The probe orientation was described in terms of the angular disorder of the filament axis about the laboratory vertical ((Theta) ) and the angle between the emission dipole and the long filament axis ((phi) ). Measurements of the prompt fluorescence from erythrosin indicated that (Theta) equals 16+/- 1 degree(s) and (phi) equals 42.1+/- 0.2 degree(s) at 20 degree(s)C while similar analyses of the delayed fluorescence gave (Theta) equals 6.4+/- 0.1 degree(s) and (Phi) equals 48.9+/- 0.1 degree(s). The probe dipole orientation ((Phi) ) was not affected by the binding of the fungal toxin phalloidin, the thin filament regulatory protein tropomyosin, or isolated myosin heads (S1) with or without nucleotide, indicating that changes in the filament anisotropy due to interaction with these ligands reflect changes in filament rotational motion. Although the angular disorder of the filaments ((Theta) ) was not influenced by ligand binding on the prompt fluorescence time scale ((tau) f is approximately 0.2 ns), it was modulated on the delayed fluorescence time scale ((tau) p is approximately 275 microsecond(s) ); the latter changes appeared to correlate with isotropic measurements of filament dynamics that indicated that phalloidin, tropomyosin, or S1 binding increased the rigidity of F-actin.

Paper Details

Date Published: 17 August 1994
PDF: 8 pages
Proc. SPIE 2137, Time-Resolved Laser Spectroscopy in Biochemistry IV, (17 August 1994); doi: 10.1117/12.182754
Show Author Affiliations
Choi-man Ng, Rutgers Univ. and Cook College (United States)
Richard D. Ludescher, Rutgers Univ. and Cook College (United States)

Published in SPIE Proceedings Vol. 2137:
Time-Resolved Laser Spectroscopy in Biochemistry IV
Joseph R. Lakowicz, Editor(s)

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