Merck
  • Home
  • Search Results
  • DeSiphering receptor core-induced and ligand-dependent conformational changes in arrestin via genetic encoded trimethylsilyl 1H-NMR probe.

DeSiphering receptor core-induced and ligand-dependent conformational changes in arrestin via genetic encoded trimethylsilyl 1H-NMR probe.

Nature communications (2020-09-27)
Qi Liu, Qing-Tao He, Xiaoxuan Lyu, Fan Yang, Zhong-Liang Zhu, Peng Xiao, Zhao Yang, Feng Zhang, Zhao-Ya Yang, Xiao-Yan Wang, Peng Sun, Qian-Wen Wang, Chang-Xiu Qu, Zheng Gong, Jing-Yu Lin, Zhen Xu, Shao-le Song, Shen-Ming Huang, Sheng-Chao Guo, Ming-Jie Han, Kong-Kai Zhu, Xin Chen, Alem W Kahsai, Kun-Hong Xiao, Wei Kong, Fa-Hui Li, Ke Ruan, Zi-Jian Li, Xiao Yu, Xiao-Gang Niu, Chang-Wen Jin, Jiangyun Wang, Jin-Peng Sun
ABSTRACT

Characterization of the dynamic conformational changes in membrane protein signaling complexes by nuclear magnetic resonance (NMR) spectroscopy remains challenging. Here we report the site-specific incorporation of 4-trimethylsilyl phenylalanine (TMSiPhe) into proteins, through genetic code expansion. Crystallographic analysis revealed structural changes that reshaped the TMSiPhe-specific amino-acyl tRNA synthetase active site to selectively accommodate the trimethylsilyl (TMSi) group. The unique up-field 1H-NMR chemical shift and the highly efficient incorporation of TMSiPhe enabled the characterization of multiple conformational states of a phospho-β2 adrenergic receptor/β-arrestin-1(β-arr1) membrane protein signaling complex, using only 5 μM protein and 20 min of spectrum accumulation time. We further showed that extracellular ligands induced conformational changes located in the polar core or ERK interaction site of β-arr1 via direct receptor transmembrane core interactions. These observations provided direct delineation and key mechanism insights that multiple receptor ligands were able to induce distinct functionally relevant conformational changes of arrestin.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Monoclonal ANTI-FLAG® M2 antibody produced in mouse, clone M2, purified immunoglobulin (Purified IgG1 subclass), buffered aqueous solution (10 mM sodium phosphate, 150 mM NaCl, pH 7.4, containing 0.02% sodium azide)
Sigma-Aldrich
Anti-Rabbit IgG (whole molecule)–Peroxidase antibody produced in goat, affinity isolated antibody, buffered aqueous solution
Sigma-Aldrich
Anti-Mouse IgG (whole molecule)–Peroxidase antibody produced in goat, affinity isolated antibody, buffered aqueous solution