This disclosure is directed to methods of conducting dynamic single-molecule fluorescence studies such as smFRET on a membrane protein which permits observation and quantification of conformational dynamics of a membrane protein. Also disclosed herein are mutant membrane proteins in which one or more mutations have been introduced for affixing a fluorophore, as well as reagents and kits containing such mutant membrane proteins for conducting dynamic single-molecule fluorescence studies. The methods and compositions disclosed herein can be used in screening for compounds that enhance or reduce the activity of a membrane protein, useful for treating diseases associated with the malfunction of the membrane protein or alterations in membrane protein conformation.