Figure S1 - FRET-based clamp binding and release assays. (a) Steady-state fluorescence emission spectra obtained with excitation at 460 nm. Donor & acceptor samples contained 500 nM fluorescein-labeled D106C delta-A in clamp loader + 4 uM D253C beta labeled with TMR. Donor only samples contained 500 nM fluorescein-labeled D106C delta-A in clamp loader + 4 uM unlabeled beta. Acceptor only samples contained 500 nM unlabeled clamp loader + 4 µM D253C beta labeled with TMR. Note that the fluorescence intensity scales are different for each. (b) Changes in acceptor emission with the addition of nucleotide and DNA primer-template. The donor contribution to observed signal (dotted green traces) was calculated by scaling the magnitude of the donor only curves in (a) to match the donor intensity observed in the presence of acceptor. Comparison with the sum of the donor contribution and the acceptor only samples (dotted blue traces) reveals that donor-dependent increases in the acceptor region of the spectra are evident in the observed signal (black traces). Fluorescence intensity is shown in arbitrary units (AU).