| Abstract: | Fluorescence energy transfer was measured by time-resolved and steady-state fluorimetry in order to investigate the spatial relationships between the nucleotide binding site of actin, the Cys-373 residue of actin, and the SH1 of myosin subfragment-1 in the rigor complex of acto-subfragment-1. N-Iodoacetyl-N'-(5-sulfo-1-naphthyl)ethylenediamine (IAEDANS) bound to the Cys-373 of actin or the fluorescent ADP analogue 1-N6-ethenoadenosine-5'-diphosphate (epsilon-ADP) bound to F-actin was used as a donor and 4-(N-(iodoacetoxy)ethyl-N-methyl)amino-7-nitrobenz-2-oxa-1,3-diazo le (IANBD) or 5-iodoacetamidofluorescein (IAF) bound to SH1 of myosin subfragment-1 was used as an acceptor. Assuming the random orientation factor, K2, to be 2/3, the distance between Cys-373 residue of actin and SH1 of myosin subfragment-1 was calculated to be about 50 A, in agreement with the values previously reported, 60 A (Takashi, R. (1969) Biochemistry 18, 5164-69) and 50 A (Trayer, H.R. and Trayer, I.P. (1983) Eur. J. Biochem. 135, 47-59). The distance between the nucleotide binding site of actin and SH1 of myosin subfragment-1 was calculated to be about 70 A or greater. |
