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

Twenty analogs of [Orn ,D-Ala ]α-factor were synthesized and assayed for their biological activities: seven analogs of [Orn ,X ]α-factor, seven analogs of [X ,D-Ala ]α-factor, five analogs of [X ,X ,D-Ala ]α-factor, and native α-factor (X = amino acids). Their biological activities (halo, gene induc...

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Bibliographic Details
Published inJournal of microbiology and biotechnology
Main Authors Hong, JinWoo, Ahn, Hee Jun, Baek, Jee Su, Hong, Eun Young, Jin, DongHoon, Khang, Yong Ho, Hong, Namjoo
Format Journal Article
LanguageEnglish
Published Korea (South) 20.09.2018
Subjects
LM102
fluorescence detector
a-factor
affinity
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Summary:Twenty analogs of [Orn ,D-Ala ]α-factor were synthesized and assayed for their biological activities: seven analogs of [Orn ,X ]α-factor, seven analogs of [X ,D-Ala ]α-factor, five analogs of [X ,X ,D-Ala ]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- vector. [Dap ,D-Ala ]α-factor with weak halo activity (10%) showed the highest receptor affinity (> 230%) and the highest gene induction activity (167%). [Arg ,D-Ala ]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 ,D-Ala ]α-factor-Edan, with high sensitivity (12,500-fold higher than the absorption-based detector [Orn ]α-factor-[Cys] ).
ISSN:1738-8872