- #FIND PEAKS ABOVE 5.00 RMSD IN COOT SOFTWARE PATCH#
- #FIND PEAKS ABOVE 5.00 RMSD IN COOT SOFTWARE FULL#
( E) Top view of the CoASH-binding site from the cytosolic side. The result is fit with the single-site-binding isotherm model with Δ H = –31.09 ± 0.97 kcal/mol and K d = 0.685 ± 0.045 μM. The background heat of control was subtracted from the heat generated during binding of CoASH to HsTMEM120A. ( D) Isothermal titration calorimetry analysis on the kinetic interactions between CoASH and HsTMEM120A. The chemical models of CoASH molecule and its fragments are shown above the corresponding peaks with m/z values 768.1232, 428.0344, and 261.1280. ( C) Mass spectrometry analysis of the small molecule extracted from the purified HsTMEM120A protein. ( B) The cryo-electron microscopy (cryo-EM) density of the ligand molecule bound to HsTMEM120A fitted with a refined structural model of CoASH. The CoASH molecules are presented as sphere models. Left, side view right, top view along the membrane normal from the intracellular side. ( A) Electrostatic potential surface presentation of HsTMEM120A dimer reveals a deep coenzyme A (CoASH)-binding cavity with an electropositive surface. The unit for the numbers labeled nearby the gradient color bar is Å. ( G) Estimation of the local resolution of the final cryo-EM map.
#FIND PEAKS ABOVE 5.00 RMSD IN COOT SOFTWARE FULL#
( F) The FSC curves between the structural model and full map as well as two half maps. ( E) Angular distribution of the particles used for the final reconstruction.
Blue, the FSC curve obtained without application of a mask orange, the FSC curve obtained with a spherical mask green and red, the FSC curve obtained with loose and tight masks around the protein density, respectively purple, the FSC curve obtained with a corrected mask. ( D) The gold standard Fourier shell correlation (GSFSC) curves for the 3D refinement of the final cryo-EM map. ( C) The flow chart of data processing, 3D classification, and refinement.
( B) Representative cryo-electron microscopy (cryo-EM) image of the HsTMEM120A-CoASH complex in nanodiscs and 2D classification images. The image of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of the peak fraction is shown on the right. ( A) Size-exclusion chromatography of the HsTMEM120A-CoASH complex reconstituted in nanodiscs. One-way analysis of variance (ANOVA) with comparison to the vector. In panels ( C), (F), and (I), each bar represents mean ± sem, and the recorded cell number is labeled above the bar. ( I) Scatter plot of the maximal stretch-induced currents.
#FIND PEAKS ABOVE 5.00 RMSD IN COOT SOFTWARE PATCH#
( H) Current-pressure curves showing stretch-induced currents under inside-out patch configuration from P1-KO-HEK cells transfected with the indicated constructs in response to the increased negative pressures. ( G) Representative stretch-induced currents from P1-KO-HEK cells transfected with the indicated constructs under the inside-out patch configuration. ( F) Scatter plot of the maximal stretch-induced currents. The MmTMEM120A group includes data from both MmTMEM120A-mCherry- and MmTMEM120A-ires-GFP-transfected cells, while the HsTMEM120A group includes data from both HsTMEM120A-mCherry- and GFP- HsTMEM120A-transfected cells. ( E) Current-pressure curves showing stretch-induced currents under cell-attached patch configuration from P1-KO-HEK cells transfected with the indicated constructs in response to the increased negative pressures. ( D) Representative stretch-induced currents from P1-KO-HEK cells transfected with the indicated constructs under the cell-attached patch configuration. ( C) Scatter plot of the maximal poking-evoked whole-cell currents. The MmTMEM120A group includes data from both MmTMEM120A-mCherry- and MmTMEM120A-ires-GFP-transfected cells.
( B) Current-displacement curves showing poking-evoked whole-cell currents from P1-KO-HEK cells transfected with the indicated constructs in response to the increased probe displacement steps. ( A) Representative poking-evoked whole-cell currents from P1-KO-HEK cells transfected with the indicated constructs.
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