Photoactivated Conformational Changes The Rhodopsin Activation Switch

Photoactivated Conformational Changes The Rhodopsin Activation Switch 

Analysis of Structural Changes from R1 Mobility and Interhelical Distances Measured by Spin—Spin Interactions 

For each of the spin-labeled sites throughout the cytoplasmic domain of rhodopsin, EPR spectra were recorded in the dark and after photoactivation to the metarhodopsin II (Mil) state, the state of rhodopsin competent for activation of transducin (Hofmann, 2000). Changes in the structure of the protein are reflected by changes in the mobility of the R1 side chains, which provide a map of the tertiary contact surfaces that rearrange during the transition to the activated form. 

Interhelical Cross-Linking Experiments and the Functional Relevance of the 

Are the helical movements detected by the spin labels functionally important If so, a molecular cross-link between the helix in question and an adjacent helix that does not move or moves in a different direction should block activation of transducin. To implement this strategy to identify functionally relevant helix motions, two different kinds of interhelical cross-links have been employed. In the first, helices are cross-linked by spontaneous or catalyzed disulfide formation between two cysteines, one engineered in each helix. In the second, cross-links are introduced by binding of Zn2+ to engineered histidine ligands, one in each helix (Elling et al., 1995). 

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V. Photoactivated Conformational Changes The Rhodopsin Activation Switch. 277... 

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