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Direct Observation of APT Protein Dynamics Using Tryptophan Fluorescence
Author(s) -
Zeheralis Chris Louis,
Johnson R. Jeremy
Publication year - 2020
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2020.34.s1.04134
Subject(s) - tryptophan , serine , chemistry , biophysics , membrane , loop (graph theory) , biochemistry , fluorescence , dynamics (music) , ligand (biochemistry) , microbiology and biotechnology , biology , enzyme , amino acid , receptor , physics , mathematics , combinatorics , acoustics , quantum mechanics
Human acyl‐protein thioesterases (APTs) catalyze the depalmitoylation of key signaling proteins attached to the plasma membrane. Catalysis was hypothesized to be regulated by the large‐scale movement of an essential dynamic loop, which transitioned the protein between active and inactive states. However, the causes for the movement of this loop are unknown. Herein, we investigated several potential factors that trigger loop movement in APTs. Specifically, we measured the effect of two irreversible serine hydrolase inhibitors and various membrane lipid models on this dynamic loop movement using intrinsic tryptophan fluorescence. Using a library of combinatorial tryptophan variants of human APT1 and APT2, we followed dynamic changes in their structure in response to shifting ligand and membrane incubations. Amongst these various conditions, the APT specific inhibitor Palmostatin B showed the largest specific effect on loop dynamics. Relating these loop dynamics to sites of modification allowed us to identify distinct regions of APTs interacting with inhibitors and with membranes. Support or Funding Information Funded by NSF MCB‐1812971