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Twenty analogs of [Orn 6 ,D-Ala 9 ]α-factor were synthesized and assayed for their biological activities: seven analogs of [Orn 6 ,X 9 ]α-factor, seven analogs of [X 6 ,D-Ala 9 ]α-factor, five analogs of [X 5 ,X 6 ,D-Ala 9 ]α-factor, and native α-factor (X = amino acids). Their biological activities (halo, gene induction, and affinity) were measured using S. cerevisiae Y7925 and LM102 and compared with those of native α-factor (100%). G protein-coupled receptor was expressed in strain LM102 containing pESC-LEU- STE2 vector. [Dap 6 ,D-Ala 9 ]α-factor with weak halo activity (10%) showed the highest receptor affinity ( 230%) and the highest gene induction activity (167%). [Arg 6 ,D-Ala 9 ]α-factor showed the highest halo activity (2,000%). The number of active binding sites per cell (about 20,000 for strain LM102) was determined using a newly-designed fluorescence-based detector, [Arg 6 ,D-Ala 9 ]α-factor-Edan, with high sensitivity (12,500-fold higher than the absorption-based detector [Orn 6 ]α-factor-[Cys] 3 ).

Development of a Highly Active Fluorescence-Based Detector for Yeast G Protein-Coupled Receptor Ste2p