Fig. 4.

Tyr223^5.58-Met257^6.40 and Tyr306^7.53-Met257^6.40 contacts in Meta II. Rows through the Met-Cε diagonal resonance are shown from 2D ¹³C DARR NMR spectra of rhodopsin (black line) and Meta II (red line) labeled with ¹³Cζ-tyrosine and ¹³Cε-methionine. The rhodopsin and Meta II rows are overlaid with the corresponding rows obtained of the Y223F (blue line) and Y306F (green line) mutants in order to determine if either of these tyrosines contact Met257^6.40 in Meta II. Upon mutation of Tyr223^5.58 or Tyr306^7.53, there is loss of intensity of the crosspeak at 156 ppm, consistent with these tyrosines having similar chemical shifts and interacting with Met257^6.40 in Meta II.