Virus:Article Title: Dynamics of the PAC1 receptor in apo, peptide agonist-bound, and G protein complexed states.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Bacterial and virus strains BL21 (DE3) Competent E.coli New England Biolabs C2527H Chemicals, peptides, and recombinant proteins Protease inhibitor cocktail Merck Cat#11697498001 Deuterium oxide Merck Cat#151882-25G ENZYMATE BEH Pepsin 2.1x30 Column Waters Cat#186007233 ACQUITY UPLC BEH C18 1.7μm 1.0x100mm Col Waters Cat#186002346 ACQUITY UPLC BEH C18 1.7μm 1.0x100mm Col Waters Cat# 186002346 PACAP38 GL Biochem N/A FLAG resin In-house N/A Lauryl maltose neopentyl glycol Anatrace Cat#NG310 ESF 921 medium Expression Systems Cat96-001-01 Apyrase New England Biosciences Cat#M0398L Benzonase Millipore Sigma Cat#71205 Flag peptide Merck Millipore Cat#F3290 Amicon Ultra Centrifugal Filter (MWCO, 100kDa) Millipore Sigma Cat#UFC910024 Amicon Ultra Centrifugal Filter (MWCO, 50kDa) Millipore Sigma Cat#UFC905008 Amicon Ultra Centrifugal Filter (MWCO, 10kDa) Millipore Sigma Cat#UFC901008 Superdex 200 10/300 GL Cytiva Cat#28990944 Cholesterol hemisuccinate Anatrace Cat#CH210 Critical commercial assays Bac-to-Bac system Thermo Fisher Cat#10359016 Deposited data HDX-MS data This paper https://www.proteomexchange.org, identifier PXD070584 Cryo-EM structure of PAC1R-Gs Piper et al. 16 PDB: 8E3X Experimental models: Cell lines High Five TM Cells Thermo Fisher Cat#B85502 Recombinant DNA HA-FLAG-3C-PAC1nR-3C-His (pfastBac dual) Piper et al. 16 N/A Gβ1/Gγ2 (pFastbac dual) Piper et al. 16 N/A Dominant negative Gαs (pFastbac) Liang et al. 30 N/A Dominant negative Gα11 (pFastbac) Josephs et al. 15 N/A Wild type Gαs (pFastbac) Piper et al. 16 N/A Nb35 Rasmussen et al. 60 N/A Software and algorithms UCSF ChimeraX 1.9 Meng et al. 61 https://www.cgl.ucsf.edu/chimerax/ Deuteros 2.0 Lau et al. 62 N/A ProteinLynx Global Server 3.0 Waters Inc. N/A (Continued on next page) Structure 34, 1–14.e1–e4, May 7, 2026 e1 .. Trichoplusia ni (High five, Thermo Fisher) insect cells and E. coli BL21 (DE3) were used in this study.
Recombinant:Article Title: Dynamics of the PAC1 receptor in apo, peptide agonist-bound, and G protein complexed states.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Bacterial and virus strains BL21 (DE3) Competent E.coli New England Biolabs C2527H Chemicals, peptides, and recombinant proteins Protease inhibitor cocktail Merck Cat#11697498001 Deuterium oxide Merck Cat#151882-25G ENZYMATE BEH Pepsin 2.1x30 Column Waters Cat#186007233 ACQUITY UPLC BEH C18 1.7μm 1.0x100mm Col Waters Cat#186002346 ACQUITY UPLC BEH C18 1.7μm 1.0x100mm Col Waters Cat# 186002346 PACAP38 GL Biochem N/A FLAG resin In-house N/A Lauryl maltose neopentyl glycol Anatrace Cat#NG310 ESF 921 medium Expression Systems Cat96-001-01 Apyrase New England Biosciences Cat#M0398L Benzonase Millipore Sigma Cat#71205 Flag peptide Merck Millipore Cat#F3290 Amicon Ultra Centrifugal Filter (MWCO, 100kDa) Millipore Sigma Cat#UFC910024 Amicon Ultra Centrifugal Filter (MWCO, 50kDa) Millipore Sigma Cat#UFC905008 Amicon Ultra Centrifugal Filter (MWCO, 10kDa) Millipore Sigma Cat#UFC901008 Superdex 200 10/300 GL Cytiva Cat#28990944 Cholesterol hemisuccinate Anatrace Cat#CH210 Critical commercial assays Bac-to-Bac system Thermo Fisher Cat#10359016 Deposited data HDX-MS data This paper https://www.proteomexchange.org, identifier PXD070584 Cryo-EM structure of PAC1R-Gs Piper et al. 16 PDB: 8E3X Experimental models: Cell lines High Five TM Cells Thermo Fisher Cat#B85502 Recombinant DNA HA-FLAG-3C-PAC1nR-3C-His (pfastBac dual) Piper et al. 16 N/A Gβ1/Gγ2 (pFastbac dual) Piper et al. 16 N/A Dominant negative Gαs (pFastbac) Liang et al. 30 N/A Dominant negative Gα11 (pFastbac) Josephs et al. 15 N/A Wild type Gαs (pFastbac) Piper et al. 16 N/A Nb35 Rasmussen et al. 60 N/A Software and algorithms UCSF ChimeraX 1.9 Meng et al. 61 https://www.cgl.ucsf.edu/chimerax/ Deuteros 2.0 Lau et al. 62 N/A ProteinLynx Global Server 3.0 Waters Inc. N/A (Continued on next page) Structure 34, 1–14.e1–e4, May 7, 2026 e1 .. Trichoplusia ni (High five, Thermo Fisher) insect cells and E. coli BL21 (DE3) were used in this study.
Article Title: Metamorphic proteins at the basis of human autophagy initiation and lipid transfer.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER 1,2-dioleoyl-sn-glycero-3phosphoethanolamine-N-(7-nitro-2-1,3benzoxadiazol-4-yl) (NBD-PE) Avanti Polar Lipids Cat#810145 1,2-dioleoyl-sn-glycero-3phosphoethanolamine-N-(lissamine rhodamine B sulfonyl) (RhoD-PE) Avanti Polar Lipids Cat#810150 1,2-dioleoyl-sn-glycero-3-phospho(1’-myo-inositol-3’-phosphate) (PI3P) Avanti Polar Lipids Cat#850150 Dodecyl-b-D-maltoside (DDM) Carl Roth Cat#69227-93-6 n-Decyl b-maltoside (DM) Glycon Biochemical Cat#D322LA Triton X-100 Merck Cat#9036-19-5 Digitonin Sigma Aldrich Cat#D141 Tween 20 Roth Cat#P1379 Amylose resin NEB Cat#E8021L Pierce Glutathione Agarose Thermo Fisher Scientific Cat#16101 Pierce Detergent Removal Resin Thermo Fisher Scientific Cat#87780 Strep-Tactin Superflow Plus Qiagen Cat#30004 HisPur Cobalt Resin Thermo Fisher Scientific Cat#89964 Natriumhypodisulfit (Dithionite) Sigma Aldrich Cat#7775-14-6 Nycodenz Alere Technologies Cat#AXS-1002424 Slide-A-Lyzer Dialysekassetten, 10 K MWCO, 12 ml Thermo Fisher Scientific Cat#66807 SERVA FastLoad 1 kb DNA Ladder SERVA Cat#39317.01 SERVA Fastload 100 bp DNA Ladder SERVA Cat#39316.01 SERVA DNA Stain G SERVA Cat#39803 All Blue Prestained Protein Standards BioRad Cat#1610373 Protein Dual Color Standards BioRad Cat#1610374 Gel Loading Dye, Purple (6X) NEB Cat#B7024S BS3 (Bis(sulfosuccinimidyl)suberat) Thermo Fisher Scientific Cat#21580 Maltose Monohydrat Merck Cat#6363-53-7 L-Glutathion Merck Cat#70-18-8 X-TREMEGENE 9 DNA Tranfection Merck Cat#6365779001 Polycarbonate Membranes 0.4mm Avanti Cat#610007 Polycarbonate Membranes 0.1mm Avanti Cat#610005 10mm Filter Supports Avanti Cat#610014 Mini-PROTEAN TGX Stain-Free Protein Gels BioRad Cat#4568034 4–15% Mini-PROTEAN TGX Precast Protein Gels BioRad Cat#4561083 96-well microplate (non-binding) Greiner Bio-one Cat#655906 0.45um Syringe Filter AMSTAT Cat#C0000629 Amicon Ultra-15 concentrator 100 kDa Milipore Cat#UFC910024 Amicon Ultra-15 concentrator 30 kDa Milipore Cat#UFC903024 T5 exonuclease Epicentre Cat#T5E4111K BSA Roth Cat#8076 DAPI Roth Cat#6335 Phusion High-Fidelity DNA Polymerase NEB Cat#M0530 Q5 High-Fidelity DNA Polymerase NEB Cat#M0491 Taq DNA Polymerase NEB Cat#M0267 Taq DNA Ligase NEB Cat#M0208 BamHI NEB Cat#R0136S (Continued on next page) e2 Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023 .. REAGENT or RESOURCE SOURCE IDENTIFIER HindIII NEB Cat#R0104S Proteinase K Sigma Alrich Cat#39450-01-6 Benzonase Merck Cat#E1014 Dpn1 NEB Cat#R0176S Critical commercial assays QIAprep Spin Miniprep Kit Qiagen Cat#27106 Plasmid Mediprep Kit Invitrogen Cat#K210015 QIAquick PCR purification Kit Qiagen Cat#28106 HiPPR Detergent Removal Spin Column Kit Thermo Fisher Scientific Cat#PI88305 Atto 647N Protein Labeling Kit Jena Bioscience Cat#FP-201-647N Alexa Fluor 488 Protein Labeling Kit Invitrogen Cat#A10235 Zeba Spin-Entsalzungss€aulen, 7 K MWCO, 0,5 ml Thermo Fisher Scientific Cat#89882 LB medium BioChemica Cat#23143289 LB-agar Roth Cat#X969.2 ESF 921 medium Expression Systems Cat96-001-01 Sf-900 medium Gibco Cat#12658-019 Bafilomycin A1 Sigma-Aldrich Cat#B1793 DPBS Gibco Cat#14190-094 EBSS Gibco Cat#24010-043 GlutaMAX -I Gibco Cat#36600-021 FuGENE 6 Promega Cat#E2691 Polybrene Sigma Alrich Cat#H9268-106; Cat#sc-134220 puromycin InvivoGen Cat#ant-pr-1 blasticidin InvivoGen Cat#ant-bl-05 PhosSTOP Roche Cat#04906837001 Deposited data Uncropped Western blot and gels images, raw microscopy images This paper, Mendeley data https://doi.org/10.17632/dyy8xfvpkf.1 Experimental models: Cell lines Sf9 insect cells Invitrogen Cat#10503433 High Five insect cells Invitrogen Cat#10747474 Oligonucleotides Oligonucleotides used in this study This study Table S4 Recombinant DNA pLIB-6x-His-MBP-ATG9 This study N/A pLIB-ATG2-StrepII This study N/A pLIB-6xHis-GFP-WIPI4 This study N/A pLIB-GST-WIPI4 This study N/A pLIB-ATG101-StrepII This study N/A pBIG-6x-His-MBP-ATG13HORMA-ATG101 This study N/A pBIG-GST-ATG13HORMA-ATG101 This study N/A pLIB-6x-His-MBP-FIP200 This study N/A pLIB-6x-His-MBP-ATG13 This study N/A pLIB-6x-His-MBP-BECN1 This study N/A pLIB-GST-ULK1 This study N/A pLIB-6xHis-mCherry-ATG14 This study N/A pLIB-6x-His-MBP-ATG13HORMA This study N/A (Continued on next page) Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023 e3 .. REAGENT or RESOURCE SOURCE IDENTIFIER pLIB-GST- ATG13HORMA This study N/A pLIB-GST- ATG13HORMA_DSB1 This study N/A pLIB-6x-His-MBP- ATG13HORMA_DSB1 This study N/A pLIB-6x-His-MBP- ATG13HORMA_DSB2 This study N/A pLIB-6x-His-MBP-ATG101 This study N/A pLIB-GST-ATG101 This study N/A pLIB-GST-ATG9C This study N/A pLIB-GST-ATG9C, P833A, P834A This study N/A pLIB-6xHis-mCherry-ATG9N This study N/A pLIB-GST-ATG9N This study N/A pLIB-6x-His-MBP-ATG9N This study N/A pLIB-6xHis-GFP-ATG14 This study N/A pLIB-BECN1 This study N/A pLIB-6x-His-MBP-ATG9core This study N/A pLIB-6x-His-MBP-ATG9core+C This study N/A pLIB-6x-His-MBP-ATG9N+core This study N/A pBIG-GST-ATG13HORMA-6xHismCherry-ATG9N This study N/A pLIB-6x-His-MBP- ATG13HORMA_DN This study N/A pLIB-6x-His-MBP- ATG13LL This study N/A Software and algorithms JalView Barton Group (University of Dundee) 2.11.1.5 PyMol Schrödinger, LLC 2.5.2 AlphaFold 2 DeepMind (Alphabet Inc.) 2 Adobe Creative Cloud Adobe Inc. 5.8.0.592 Graphpad PRISM GraphPad Software Inc. 9.0.0 ImageJ NIH, USA 1.53t Image Studio Lite LI-COR Biosciences 5.2 ChimeraX 1.4 UCSF 1.4 Snapgene GSL Biotech 5.3.2 Fiji Schindelin et al.68 1.53 Microsoft Office Microsoft Corporation 16.64 Others MBPTrap HP (1 ml; 5 ml) Cytiva Cat#29048641; Cat#28918778 HiTrap TALON crude (1 ml; 5ml) Cytiva Cat#29048565; Cat#28953766 GSTrap HP (1 ml; 5 ml) Cytiva Cat#17528101; Cat#17528201 StrepTrap HP (5ml) Cytiva Cat#28907546 HiLoad 16/600 Superdex 75 pg Cytiva Cat#28989333 HiLoad 16/600 Superdex 200 pg Cytiva Cat#28989335 HiLoad 16/600 Superose 6 pg Cytiva Cat#29323952 Superdex 75 Increase 10/300 GL Cytiva Cat#29148721 Superdex 200 Increase 5/150 GL Cytiva Cat#28990945 Superose 6 Increase 10/300 GL Cytiva Cat#29091596 Superose 6 Increase 5/150 GL Cytiva Cat#29091597 RESOURCE Q, 6 ml Cytiva Cat#17117901 ProLong Glass Antifade Mountant Thermo Fisher Scientific Cat#P36980 e4 Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023
Article Title: Activation and closed-state inactivation mechanisms of the human voltage-gated K V 4 channel complexes.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Bacterial strains Turbo competent cells New England Biolabs Cat#C2984H DH10Bac competent cells Thermo Fisher Scientific Cat#10361012 Chemicals, peptides, and recombinant proteins Protease inhibitor cocktail MedChemExpress Cat#HY-K0010 Lauryl maltose neopentyl glycol (LMNG) Anatrace Cat#NG310 n-Dodecyl-b-D-Maltopyranoside (DDM) Anatrace Cat#D310S Glyco-Diosgenin (GDN) Anatrace Cat#GDN101 Cholesteryl hemisuccinate tris salt Anatrace Cat#CH210 ESF 921 Medium Expression Systems Cat96-001-01 Freestyle 293 expression medium Gibco Cat#12338018 DMEM/F-12 medium Gibco Cat#11330-032 Penicillin-streptomycin solution, 100x Corning Cat#30-002-CI Fetal bovine serum Gibco Cat#16140089 Lipofectamine 2000 reagent Invitrogen Cat#11668019 TransIT-Insect transfection reagent Mirus Cat#MIR6100 1,1-Methanediyl Bismethanethiosulfonate (MTS-1-MTS) Santa Cruz Biotech Cat#SC-208706 1,4-Butanediyl Bismethanethiosulfonate (MTS-4-MTS) Santa Cruz Biotech Cat#SC-208789 Deposited data KV4.2/KChIP2 channel complex, open state This study PDB: 7UK5; EMDB: EMD-26575 KV4.2/KChIP2 channel complex, inactivated state, class 1 This study PDB: 7UKC; EMDB: EMD-26576 KV4.2/KChIP2 channel complex, inactivated state, class 2 This study PDB: 7UKD; EMDB: EMD-26577 KV4.2/KChIP2 channel complex, intermediate state This study PDB: 7UKE; EMDB: EMD-26578 KV4.2/KChIP2 channel complex, putative resting state This study PDB: 7UKF; EMDB: EMD-26579 KV4.2/KChIP2/DPP6 channel complex, open state, transmembrane region This study PDB: 7UKG; EMDB: EMD-26580 KV4.2/KChIP2/DPP6 channel complex, open state, intracellular region This study PDB: 7UKH; EMDB: EMD-26581 Experimental models: Cell lines HEK293 ATCC Cat#CRL-1573 Sf9 ATCC Cat#CRL-1711 HEK293S GnTI– ATCC Cat#CRL-3022 Recombinant DNA pEG-BacMam Human Kv4.2EM This study N/A pEG-BacMam Human KChIP2 This study N/A pEG-BacMam Human DPP6 This study N/A Software and algorithms Serial EM Mastronarde (2005) http://bio3d.colorado.edu/SerialEM RELION Zivanov et al. (2018) https://github.com/3dem/relion (Continued on next page) Molecular Cell 82, 2427–2442.e1–e4, July 7, 2022 e1 .. REAGENT or RESOURCE SOURCE IDENTIFIER cryoSPARC Punjani et al. (2017) https://cryosparc.com/ Coot Emsley et al. (2010) http://www2.mrc-lmb.cam.ac.uk/personal/ pemsley/coot Phenix Afonine et al. (2018) https://www.phenix-online.org MolProbity Chen et al. (2010) http://molprobity.biochem.duke.edu Chimera Pettersen et al. (2004) https://www.cgl.ucsf.edu/chimera PyMOL Schrodinger http://www.pymol.org pCLAMP Molecular Devices https://www.moleculardevices.com/ Anaconda Anaconda https://www.anaconda.com/ Other CNBR-activated Sepharose beads Cytiva Cat#17043001 Superose 6 Increase 10/300 GL SEC column Cytiva Cat#29091596 R1.2/1.3 400 mesh UltrAuFoil gold grids Quantifoil Cat#Q350AR13A
Protease Inhibitor:Article Title: Dynamics of the PAC1 receptor in apo, peptide agonist-bound, and G protein complexed states.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Bacterial and virus strains BL21 (DE3) Competent E.coli New England Biolabs C2527H Chemicals, peptides, and recombinant proteins Protease inhibitor cocktail Merck Cat#11697498001 Deuterium oxide Merck Cat#151882-25G ENZYMATE BEH Pepsin 2.1x30 Column Waters Cat#186007233 ACQUITY UPLC BEH C18 1.7μm 1.0x100mm Col Waters Cat#186002346 ACQUITY UPLC BEH C18 1.7μm 1.0x100mm Col Waters Cat# 186002346 PACAP38 GL Biochem N/A FLAG resin In-house N/A Lauryl maltose neopentyl glycol Anatrace Cat#NG310 ESF 921 medium Expression Systems Cat96-001-01 Apyrase New England Biosciences Cat#M0398L Benzonase Millipore Sigma Cat#71205 Flag peptide Merck Millipore Cat#F3290 Amicon Ultra Centrifugal Filter (MWCO, 100kDa) Millipore Sigma Cat#UFC910024 Amicon Ultra Centrifugal Filter (MWCO, 50kDa) Millipore Sigma Cat#UFC905008 Amicon Ultra Centrifugal Filter (MWCO, 10kDa) Millipore Sigma Cat#UFC901008 Superdex 200 10/300 GL Cytiva Cat#28990944 Cholesterol hemisuccinate Anatrace Cat#CH210 Critical commercial assays Bac-to-Bac system Thermo Fisher Cat#10359016 Deposited data HDX-MS data This paper https://www.proteomexchange.org, identifier PXD070584 Cryo-EM structure of PAC1R-Gs Piper et al. 16 PDB: 8E3X Experimental models: Cell lines High Five TM Cells Thermo Fisher Cat#B85502 Recombinant DNA HA-FLAG-3C-PAC1nR-3C-His (pfastBac dual) Piper et al. 16 N/A Gβ1/Gγ2 (pFastbac dual) Piper et al. 16 N/A Dominant negative Gαs (pFastbac) Liang et al. 30 N/A Dominant negative Gα11 (pFastbac) Josephs et al. 15 N/A Wild type Gαs (pFastbac) Piper et al. 16 N/A Nb35 Rasmussen et al. 60 N/A Software and algorithms UCSF ChimeraX 1.9 Meng et al. 61 https://www.cgl.ucsf.edu/chimerax/ Deuteros 2.0 Lau et al. 62 N/A ProteinLynx Global Server 3.0 Waters Inc. N/A (Continued on next page) Structure 34, 1–14.e1–e4, May 7, 2026 e1 .. Trichoplusia ni (High five, Thermo Fisher) insect cells and E. coli BL21 (DE3) were used in this study.
Article Title: Activation and closed-state inactivation mechanisms of the human voltage-gated K V 4 channel complexes.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Bacterial strains Turbo competent cells New England Biolabs Cat#C2984H DH10Bac competent cells Thermo Fisher Scientific Cat#10361012 Chemicals, peptides, and recombinant proteins Protease inhibitor cocktail MedChemExpress Cat#HY-K0010 Lauryl maltose neopentyl glycol (LMNG) Anatrace Cat#NG310 n-Dodecyl-b-D-Maltopyranoside (DDM) Anatrace Cat#D310S Glyco-Diosgenin (GDN) Anatrace Cat#GDN101 Cholesteryl hemisuccinate tris salt Anatrace Cat#CH210 ESF 921 Medium Expression Systems Cat96-001-01 Freestyle 293 expression medium Gibco Cat#12338018 DMEM/F-12 medium Gibco Cat#11330-032 Penicillin-streptomycin solution, 100x Corning Cat#30-002-CI Fetal bovine serum Gibco Cat#16140089 Lipofectamine 2000 reagent Invitrogen Cat#11668019 TransIT-Insect transfection reagent Mirus Cat#MIR6100 1,1-Methanediyl Bismethanethiosulfonate (MTS-1-MTS) Santa Cruz Biotech Cat#SC-208706 1,4-Butanediyl Bismethanethiosulfonate (MTS-4-MTS) Santa Cruz Biotech Cat#SC-208789 Deposited data KV4.2/KChIP2 channel complex, open state This study PDB: 7UK5; EMDB: EMD-26575 KV4.2/KChIP2 channel complex, inactivated state, class 1 This study PDB: 7UKC; EMDB: EMD-26576 KV4.2/KChIP2 channel complex, inactivated state, class 2 This study PDB: 7UKD; EMDB: EMD-26577 KV4.2/KChIP2 channel complex, intermediate state This study PDB: 7UKE; EMDB: EMD-26578 KV4.2/KChIP2 channel complex, putative resting state This study PDB: 7UKF; EMDB: EMD-26579 KV4.2/KChIP2/DPP6 channel complex, open state, transmembrane region This study PDB: 7UKG; EMDB: EMD-26580 KV4.2/KChIP2/DPP6 channel complex, open state, intracellular region This study PDB: 7UKH; EMDB: EMD-26581 Experimental models: Cell lines HEK293 ATCC Cat#CRL-1573 Sf9 ATCC Cat#CRL-1711 HEK293S GnTI– ATCC Cat#CRL-3022 Recombinant DNA pEG-BacMam Human Kv4.2EM This study N/A pEG-BacMam Human KChIP2 This study N/A pEG-BacMam Human DPP6 This study N/A Software and algorithms Serial EM Mastronarde (2005) http://bio3d.colorado.edu/SerialEM RELION Zivanov et al. (2018) https://github.com/3dem/relion (Continued on next page) Molecular Cell 82, 2427–2442.e1–e4, July 7, 2022 e1 .. REAGENT or RESOURCE SOURCE IDENTIFIER cryoSPARC Punjani et al. (2017) https://cryosparc.com/ Coot Emsley et al. (2010) http://www2.mrc-lmb.cam.ac.uk/personal/ pemsley/coot Phenix Afonine et al. (2018) https://www.phenix-online.org MolProbity Chen et al. (2010) http://molprobity.biochem.duke.edu Chimera Pettersen et al. (2004) https://www.cgl.ucsf.edu/chimera PyMOL Schrodinger http://www.pymol.org pCLAMP Molecular Devices https://www.moleculardevices.com/ Anaconda Anaconda https://www.anaconda.com/ Other CNBR-activated Sepharose beads Cytiva Cat#17043001 Superose 6 Increase 10/300 GL SEC column Cytiva Cat#29091596 R1.2/1.3 400 mesh UltrAuFoil gold grids Quantifoil Cat#Q350AR13A
Structural Proteomics:Article Title: Dynamics of the PAC1 receptor in apo, peptide agonist-bound, and G protein complexed states.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Bacterial and virus strains BL21 (DE3) Competent E.coli New England Biolabs C2527H Chemicals, peptides, and recombinant proteins Protease inhibitor cocktail Merck Cat#11697498001 Deuterium oxide Merck Cat#151882-25G ENZYMATE BEH Pepsin 2.1x30 Column Waters Cat#186007233 ACQUITY UPLC BEH C18 1.7μm 1.0x100mm Col Waters Cat#186002346 ACQUITY UPLC BEH C18 1.7μm 1.0x100mm Col Waters Cat# 186002346 PACAP38 GL Biochem N/A FLAG resin In-house N/A Lauryl maltose neopentyl glycol Anatrace Cat#NG310 ESF 921 medium Expression Systems Cat96-001-01 Apyrase New England Biosciences Cat#M0398L Benzonase Millipore Sigma Cat#71205 Flag peptide Merck Millipore Cat#F3290 Amicon Ultra Centrifugal Filter (MWCO, 100kDa) Millipore Sigma Cat#UFC910024 Amicon Ultra Centrifugal Filter (MWCO, 50kDa) Millipore Sigma Cat#UFC905008 Amicon Ultra Centrifugal Filter (MWCO, 10kDa) Millipore Sigma Cat#UFC901008 Superdex 200 10/300 GL Cytiva Cat#28990944 Cholesterol hemisuccinate Anatrace Cat#CH210 Critical commercial assays Bac-to-Bac system Thermo Fisher Cat#10359016 Deposited data HDX-MS data This paper https://www.proteomexchange.org, identifier PXD070584 Cryo-EM structure of PAC1R-Gs Piper et al. 16 PDB: 8E3X Experimental models: Cell lines High Five TM Cells Thermo Fisher Cat#B85502 Recombinant DNA HA-FLAG-3C-PAC1nR-3C-His (pfastBac dual) Piper et al. 16 N/A Gβ1/Gγ2 (pFastbac dual) Piper et al. 16 N/A Dominant negative Gαs (pFastbac) Liang et al. 30 N/A Dominant negative Gα11 (pFastbac) Josephs et al. 15 N/A Wild type Gαs (pFastbac) Piper et al. 16 N/A Nb35 Rasmussen et al. 60 N/A Software and algorithms UCSF ChimeraX 1.9 Meng et al. 61 https://www.cgl.ucsf.edu/chimerax/ Deuteros 2.0 Lau et al. 62 N/A ProteinLynx Global Server 3.0 Waters Inc. N/A (Continued on next page) Structure 34, 1–14.e1–e4, May 7, 2026 e1 .. Trichoplusia ni (High five, Thermo Fisher) insect cells and E. coli BL21 (DE3) were used in this study.
Cryo-EM Sample Prep:Article Title: Dynamics of the PAC1 receptor in apo, peptide agonist-bound, and G protein complexed states.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Bacterial and virus strains BL21 (DE3) Competent E.coli New England Biolabs C2527H Chemicals, peptides, and recombinant proteins Protease inhibitor cocktail Merck Cat#11697498001 Deuterium oxide Merck Cat#151882-25G ENZYMATE BEH Pepsin 2.1x30 Column Waters Cat#186007233 ACQUITY UPLC BEH C18 1.7μm 1.0x100mm Col Waters Cat#186002346 ACQUITY UPLC BEH C18 1.7μm 1.0x100mm Col Waters Cat# 186002346 PACAP38 GL Biochem N/A FLAG resin In-house N/A Lauryl maltose neopentyl glycol Anatrace Cat#NG310 ESF 921 medium Expression Systems Cat96-001-01 Apyrase New England Biosciences Cat#M0398L Benzonase Millipore Sigma Cat#71205 Flag peptide Merck Millipore Cat#F3290 Amicon Ultra Centrifugal Filter (MWCO, 100kDa) Millipore Sigma Cat#UFC910024 Amicon Ultra Centrifugal Filter (MWCO, 50kDa) Millipore Sigma Cat#UFC905008 Amicon Ultra Centrifugal Filter (MWCO, 10kDa) Millipore Sigma Cat#UFC901008 Superdex 200 10/300 GL Cytiva Cat#28990944 Cholesterol hemisuccinate Anatrace Cat#CH210 Critical commercial assays Bac-to-Bac system Thermo Fisher Cat#10359016 Deposited data HDX-MS data This paper https://www.proteomexchange.org, identifier PXD070584 Cryo-EM structure of PAC1R-Gs Piper et al. 16 PDB: 8E3X Experimental models: Cell lines High Five TM Cells Thermo Fisher Cat#B85502 Recombinant DNA HA-FLAG-3C-PAC1nR-3C-His (pfastBac dual) Piper et al. 16 N/A Gβ1/Gγ2 (pFastbac dual) Piper et al. 16 N/A Dominant negative Gαs (pFastbac) Liang et al. 30 N/A Dominant negative Gα11 (pFastbac) Josephs et al. 15 N/A Wild type Gαs (pFastbac) Piper et al. 16 N/A Nb35 Rasmussen et al. 60 N/A Software and algorithms UCSF ChimeraX 1.9 Meng et al. 61 https://www.cgl.ucsf.edu/chimerax/ Deuteros 2.0 Lau et al. 62 N/A ProteinLynx Global Server 3.0 Waters Inc. N/A (Continued on next page) Structure 34, 1–14.e1–e4, May 7, 2026 e1 .. Trichoplusia ni (High five, Thermo Fisher) insect cells and E. coli BL21 (DE3) were used in this study.
Dominant Negative Mutation:Article Title: Dynamics of the PAC1 receptor in apo, peptide agonist-bound, and G protein complexed states.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Bacterial and virus strains BL21 (DE3) Competent E.coli New England Biolabs C2527H Chemicals, peptides, and recombinant proteins Protease inhibitor cocktail Merck Cat#11697498001 Deuterium oxide Merck Cat#151882-25G ENZYMATE BEH Pepsin 2.1x30 Column Waters Cat#186007233 ACQUITY UPLC BEH C18 1.7μm 1.0x100mm Col Waters Cat#186002346 ACQUITY UPLC BEH C18 1.7μm 1.0x100mm Col Waters Cat# 186002346 PACAP38 GL Biochem N/A FLAG resin In-house N/A Lauryl maltose neopentyl glycol Anatrace Cat#NG310 ESF 921 medium Expression Systems Cat96-001-01 Apyrase New England Biosciences Cat#M0398L Benzonase Millipore Sigma Cat#71205 Flag peptide Merck Millipore Cat#F3290 Amicon Ultra Centrifugal Filter (MWCO, 100kDa) Millipore Sigma Cat#UFC910024 Amicon Ultra Centrifugal Filter (MWCO, 50kDa) Millipore Sigma Cat#UFC905008 Amicon Ultra Centrifugal Filter (MWCO, 10kDa) Millipore Sigma Cat#UFC901008 Superdex 200 10/300 GL Cytiva Cat#28990944 Cholesterol hemisuccinate Anatrace Cat#CH210 Critical commercial assays Bac-to-Bac system Thermo Fisher Cat#10359016 Deposited data HDX-MS data This paper https://www.proteomexchange.org, identifier PXD070584 Cryo-EM structure of PAC1R-Gs Piper et al. 16 PDB: 8E3X Experimental models: Cell lines High Five TM Cells Thermo Fisher Cat#B85502 Recombinant DNA HA-FLAG-3C-PAC1nR-3C-His (pfastBac dual) Piper et al. 16 N/A Gβ1/Gγ2 (pFastbac dual) Piper et al. 16 N/A Dominant negative Gαs (pFastbac) Liang et al. 30 N/A Dominant negative Gα11 (pFastbac) Josephs et al. 15 N/A Wild type Gαs (pFastbac) Piper et al. 16 N/A Nb35 Rasmussen et al. 60 N/A Software and algorithms UCSF ChimeraX 1.9 Meng et al. 61 https://www.cgl.ucsf.edu/chimerax/ Deuteros 2.0 Lau et al. 62 N/A ProteinLynx Global Server 3.0 Waters Inc. N/A (Continued on next page) Structure 34, 1–14.e1–e4, May 7, 2026 e1 .. Trichoplusia ni (High five, Thermo Fisher) insect cells and E. coli BL21 (DE3) were used in this study.
Software:Article Title: Dynamics of the PAC1 receptor in apo, peptide agonist-bound, and G protein complexed states.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Bacterial and virus strains BL21 (DE3) Competent E.coli New England Biolabs C2527H Chemicals, peptides, and recombinant proteins Protease inhibitor cocktail Merck Cat#11697498001 Deuterium oxide Merck Cat#151882-25G ENZYMATE BEH Pepsin 2.1x30 Column Waters Cat#186007233 ACQUITY UPLC BEH C18 1.7μm 1.0x100mm Col Waters Cat#186002346 ACQUITY UPLC BEH C18 1.7μm 1.0x100mm Col Waters Cat# 186002346 PACAP38 GL Biochem N/A FLAG resin In-house N/A Lauryl maltose neopentyl glycol Anatrace Cat#NG310 ESF 921 medium Expression Systems Cat96-001-01 Apyrase New England Biosciences Cat#M0398L Benzonase Millipore Sigma Cat#71205 Flag peptide Merck Millipore Cat#F3290 Amicon Ultra Centrifugal Filter (MWCO, 100kDa) Millipore Sigma Cat#UFC910024 Amicon Ultra Centrifugal Filter (MWCO, 50kDa) Millipore Sigma Cat#UFC905008 Amicon Ultra Centrifugal Filter (MWCO, 10kDa) Millipore Sigma Cat#UFC901008 Superdex 200 10/300 GL Cytiva Cat#28990944 Cholesterol hemisuccinate Anatrace Cat#CH210 Critical commercial assays Bac-to-Bac system Thermo Fisher Cat#10359016 Deposited data HDX-MS data This paper https://www.proteomexchange.org, identifier PXD070584 Cryo-EM structure of PAC1R-Gs Piper et al. 16 PDB: 8E3X Experimental models: Cell lines High Five TM Cells Thermo Fisher Cat#B85502 Recombinant DNA HA-FLAG-3C-PAC1nR-3C-His (pfastBac dual) Piper et al. 16 N/A Gβ1/Gγ2 (pFastbac dual) Piper et al. 16 N/A Dominant negative Gαs (pFastbac) Liang et al. 30 N/A Dominant negative Gα11 (pFastbac) Josephs et al. 15 N/A Wild type Gαs (pFastbac) Piper et al. 16 N/A Nb35 Rasmussen et al. 60 N/A Software and algorithms UCSF ChimeraX 1.9 Meng et al. 61 https://www.cgl.ucsf.edu/chimerax/ Deuteros 2.0 Lau et al. 62 N/A ProteinLynx Global Server 3.0 Waters Inc. N/A (Continued on next page) Structure 34, 1–14.e1–e4, May 7, 2026 e1 .. Trichoplusia ni (High five, Thermo Fisher) insect cells and E. coli BL21 (DE3) were used in this study.
Article Title: Activation and closed-state inactivation mechanisms of the human voltage-gated K V 4 channel complexes.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Bacterial strains Turbo competent cells New England Biolabs Cat#C2984H DH10Bac competent cells Thermo Fisher Scientific Cat#10361012 Chemicals, peptides, and recombinant proteins Protease inhibitor cocktail MedChemExpress Cat#HY-K0010 Lauryl maltose neopentyl glycol (LMNG) Anatrace Cat#NG310 n-Dodecyl-b-D-Maltopyranoside (DDM) Anatrace Cat#D310S Glyco-Diosgenin (GDN) Anatrace Cat#GDN101 Cholesteryl hemisuccinate tris salt Anatrace Cat#CH210 ESF 921 Medium Expression Systems Cat96-001-01 Freestyle 293 expression medium Gibco Cat#12338018 DMEM/F-12 medium Gibco Cat#11330-032 Penicillin-streptomycin solution, 100x Corning Cat#30-002-CI Fetal bovine serum Gibco Cat#16140089 Lipofectamine 2000 reagent Invitrogen Cat#11668019 TransIT-Insect transfection reagent Mirus Cat#MIR6100 1,1-Methanediyl Bismethanethiosulfonate (MTS-1-MTS) Santa Cruz Biotech Cat#SC-208706 1,4-Butanediyl Bismethanethiosulfonate (MTS-4-MTS) Santa Cruz Biotech Cat#SC-208789 Deposited data KV4.2/KChIP2 channel complex, open state This study PDB: 7UK5; EMDB: EMD-26575 KV4.2/KChIP2 channel complex, inactivated state, class 1 This study PDB: 7UKC; EMDB: EMD-26576 KV4.2/KChIP2 channel complex, inactivated state, class 2 This study PDB: 7UKD; EMDB: EMD-26577 KV4.2/KChIP2 channel complex, intermediate state This study PDB: 7UKE; EMDB: EMD-26578 KV4.2/KChIP2 channel complex, putative resting state This study PDB: 7UKF; EMDB: EMD-26579 KV4.2/KChIP2/DPP6 channel complex, open state, transmembrane region This study PDB: 7UKG; EMDB: EMD-26580 KV4.2/KChIP2/DPP6 channel complex, open state, intracellular region This study PDB: 7UKH; EMDB: EMD-26581 Experimental models: Cell lines HEK293 ATCC Cat#CRL-1573 Sf9 ATCC Cat#CRL-1711 HEK293S GnTI– ATCC Cat#CRL-3022 Recombinant DNA pEG-BacMam Human Kv4.2EM This study N/A pEG-BacMam Human KChIP2 This study N/A pEG-BacMam Human DPP6 This study N/A Software and algorithms Serial EM Mastronarde (2005) http://bio3d.colorado.edu/SerialEM RELION Zivanov et al. (2018) https://github.com/3dem/relion (Continued on next page) Molecular Cell 82, 2427–2442.e1–e4, July 7, 2022 e1 .. REAGENT or RESOURCE SOURCE IDENTIFIER cryoSPARC Punjani et al. (2017) https://cryosparc.com/ Coot Emsley et al. (2010) http://www2.mrc-lmb.cam.ac.uk/personal/ pemsley/coot Phenix Afonine et al. (2018) https://www.phenix-online.org MolProbity Chen et al. (2010) http://molprobity.biochem.duke.edu Chimera Pettersen et al. (2004) https://www.cgl.ucsf.edu/chimera PyMOL Schrodinger http://www.pymol.org pCLAMP Molecular Devices https://www.moleculardevices.com/ Anaconda Anaconda https://www.anaconda.com/ Other CNBR-activated Sepharose beads Cytiva Cat#17043001 Superose 6 Increase 10/300 GL SEC column Cytiva Cat#29091596 R1.2/1.3 400 mesh UltrAuFoil gold grids Quantifoil Cat#Q350AR13A
Plasmid Preparation:Article Title: Metamorphic proteins at the basis of human autophagy initiation and lipid transfer.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER 1,2-dioleoyl-sn-glycero-3phosphoethanolamine-N-(7-nitro-2-1,3benzoxadiazol-4-yl) (NBD-PE) Avanti Polar Lipids Cat#810145 1,2-dioleoyl-sn-glycero-3phosphoethanolamine-N-(lissamine rhodamine B sulfonyl) (RhoD-PE) Avanti Polar Lipids Cat#810150 1,2-dioleoyl-sn-glycero-3-phospho(1’-myo-inositol-3’-phosphate) (PI3P) Avanti Polar Lipids Cat#850150 Dodecyl-b-D-maltoside (DDM) Carl Roth Cat#69227-93-6 n-Decyl b-maltoside (DM) Glycon Biochemical Cat#D322LA Triton X-100 Merck Cat#9036-19-5 Digitonin Sigma Aldrich Cat#D141 Tween 20 Roth Cat#P1379 Amylose resin NEB Cat#E8021L Pierce Glutathione Agarose Thermo Fisher Scientific Cat#16101 Pierce Detergent Removal Resin Thermo Fisher Scientific Cat#87780 Strep-Tactin Superflow Plus Qiagen Cat#30004 HisPur Cobalt Resin Thermo Fisher Scientific Cat#89964 Natriumhypodisulfit (Dithionite) Sigma Aldrich Cat#7775-14-6 Nycodenz Alere Technologies Cat#AXS-1002424 Slide-A-Lyzer Dialysekassetten, 10 K MWCO, 12 ml Thermo Fisher Scientific Cat#66807 SERVA FastLoad 1 kb DNA Ladder SERVA Cat#39317.01 SERVA Fastload 100 bp DNA Ladder SERVA Cat#39316.01 SERVA DNA Stain G SERVA Cat#39803 All Blue Prestained Protein Standards BioRad Cat#1610373 Protein Dual Color Standards BioRad Cat#1610374 Gel Loading Dye, Purple (6X) NEB Cat#B7024S BS3 (Bis(sulfosuccinimidyl)suberat) Thermo Fisher Scientific Cat#21580 Maltose Monohydrat Merck Cat#6363-53-7 L-Glutathion Merck Cat#70-18-8 X-TREMEGENE 9 DNA Tranfection Merck Cat#6365779001 Polycarbonate Membranes 0.4mm Avanti Cat#610007 Polycarbonate Membranes 0.1mm Avanti Cat#610005 10mm Filter Supports Avanti Cat#610014 Mini-PROTEAN TGX Stain-Free Protein Gels BioRad Cat#4568034 4–15% Mini-PROTEAN TGX Precast Protein Gels BioRad Cat#4561083 96-well microplate (non-binding) Greiner Bio-one Cat#655906 0.45um Syringe Filter AMSTAT Cat#C0000629 Amicon Ultra-15 concentrator 100 kDa Milipore Cat#UFC910024 Amicon Ultra-15 concentrator 30 kDa Milipore Cat#UFC903024 T5 exonuclease Epicentre Cat#T5E4111K BSA Roth Cat#8076 DAPI Roth Cat#6335 Phusion High-Fidelity DNA Polymerase NEB Cat#M0530 Q5 High-Fidelity DNA Polymerase NEB Cat#M0491 Taq DNA Polymerase NEB Cat#M0267 Taq DNA Ligase NEB Cat#M0208 BamHI NEB Cat#R0136S (Continued on next page) e2 Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023 .. REAGENT or RESOURCE SOURCE IDENTIFIER HindIII NEB Cat#R0104S Proteinase K Sigma Alrich Cat#39450-01-6 Benzonase Merck Cat#E1014 Dpn1 NEB Cat#R0176S Critical commercial assays QIAprep Spin Miniprep Kit Qiagen Cat#27106 Plasmid Mediprep Kit Invitrogen Cat#K210015 QIAquick PCR purification Kit Qiagen Cat#28106 HiPPR Detergent Removal Spin Column Kit Thermo Fisher Scientific Cat#PI88305 Atto 647N Protein Labeling Kit Jena Bioscience Cat#FP-201-647N Alexa Fluor 488 Protein Labeling Kit Invitrogen Cat#A10235 Zeba Spin-Entsalzungss€aulen, 7 K MWCO, 0,5 ml Thermo Fisher Scientific Cat#89882 LB medium BioChemica Cat#23143289 LB-agar Roth Cat#X969.2 ESF 921 medium Expression Systems Cat96-001-01 Sf-900 medium Gibco Cat#12658-019 Bafilomycin A1 Sigma-Aldrich Cat#B1793 DPBS Gibco Cat#14190-094 EBSS Gibco Cat#24010-043 GlutaMAX -I Gibco Cat#36600-021 FuGENE 6 Promega Cat#E2691 Polybrene Sigma Alrich Cat#H9268-106; Cat#sc-134220 puromycin InvivoGen Cat#ant-pr-1 blasticidin InvivoGen Cat#ant-bl-05 PhosSTOP Roche Cat#04906837001 Deposited data Uncropped Western blot and gels images, raw microscopy images This paper, Mendeley data https://doi.org/10.17632/dyy8xfvpkf.1 Experimental models: Cell lines Sf9 insect cells Invitrogen Cat#10503433 High Five insect cells Invitrogen Cat#10747474 Oligonucleotides Oligonucleotides used in this study This study Table S4 Recombinant DNA pLIB-6x-His-MBP-ATG9 This study N/A pLIB-ATG2-StrepII This study N/A pLIB-6xHis-GFP-WIPI4 This study N/A pLIB-GST-WIPI4 This study N/A pLIB-ATG101-StrepII This study N/A pBIG-6x-His-MBP-ATG13HORMA-ATG101 This study N/A pBIG-GST-ATG13HORMA-ATG101 This study N/A pLIB-6x-His-MBP-FIP200 This study N/A pLIB-6x-His-MBP-ATG13 This study N/A pLIB-6x-His-MBP-BECN1 This study N/A pLIB-GST-ULK1 This study N/A pLIB-6xHis-mCherry-ATG14 This study N/A pLIB-6x-His-MBP-ATG13HORMA This study N/A (Continued on next page) Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023 e3 .. REAGENT or RESOURCE SOURCE IDENTIFIER pLIB-GST- ATG13HORMA This study N/A pLIB-GST- ATG13HORMA_DSB1 This study N/A pLIB-6x-His-MBP- ATG13HORMA_DSB1 This study N/A pLIB-6x-His-MBP- ATG13HORMA_DSB2 This study N/A pLIB-6x-His-MBP-ATG101 This study N/A pLIB-GST-ATG101 This study N/A pLIB-GST-ATG9C This study N/A pLIB-GST-ATG9C, P833A, P834A This study N/A pLIB-6xHis-mCherry-ATG9N This study N/A pLIB-GST-ATG9N This study N/A pLIB-6x-His-MBP-ATG9N This study N/A pLIB-6xHis-GFP-ATG14 This study N/A pLIB-BECN1 This study N/A pLIB-6x-His-MBP-ATG9core This study N/A pLIB-6x-His-MBP-ATG9core+C This study N/A pLIB-6x-His-MBP-ATG9N+core This study N/A pBIG-GST-ATG13HORMA-6xHismCherry-ATG9N This study N/A pLIB-6x-His-MBP- ATG13HORMA_DN This study N/A pLIB-6x-His-MBP- ATG13LL This study N/A Software and algorithms JalView Barton Group (University of Dundee) 2.11.1.5 PyMol Schrödinger, LLC 2.5.2 AlphaFold 2 DeepMind (Alphabet Inc.) 2 Adobe Creative Cloud Adobe Inc. 5.8.0.592 Graphpad PRISM GraphPad Software Inc. 9.0.0 ImageJ NIH, USA 1.53t Image Studio Lite LI-COR Biosciences 5.2 ChimeraX 1.4 UCSF 1.4 Snapgene GSL Biotech 5.3.2 Fiji Schindelin et al.68 1.53 Microsoft Office Microsoft Corporation 16.64 Others MBPTrap HP (1 ml; 5 ml) Cytiva Cat#29048641; Cat#28918778 HiTrap TALON crude (1 ml; 5ml) Cytiva Cat#29048565; Cat#28953766 GSTrap HP (1 ml; 5 ml) Cytiva Cat#17528101; Cat#17528201 StrepTrap HP (5ml) Cytiva Cat#28907546 HiLoad 16/600 Superdex 75 pg Cytiva Cat#28989333 HiLoad 16/600 Superdex 200 pg Cytiva Cat#28989335 HiLoad 16/600 Superose 6 pg Cytiva Cat#29323952 Superdex 75 Increase 10/300 GL Cytiva Cat#29148721 Superdex 200 Increase 5/150 GL Cytiva Cat#28990945 Superose 6 Increase 10/300 GL Cytiva Cat#29091596 Superose 6 Increase 5/150 GL Cytiva Cat#29091597 RESOURCE Q, 6 ml Cytiva Cat#17117901 ProLong Glass Antifade Mountant Thermo Fisher Scientific Cat#P36980 e4 Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023
Polymerase Chain Reaction:Article Title: Metamorphic proteins at the basis of human autophagy initiation and lipid transfer.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER 1,2-dioleoyl-sn-glycero-3phosphoethanolamine-N-(7-nitro-2-1,3benzoxadiazol-4-yl) (NBD-PE) Avanti Polar Lipids Cat#810145 1,2-dioleoyl-sn-glycero-3phosphoethanolamine-N-(lissamine rhodamine B sulfonyl) (RhoD-PE) Avanti Polar Lipids Cat#810150 1,2-dioleoyl-sn-glycero-3-phospho(1’-myo-inositol-3’-phosphate) (PI3P) Avanti Polar Lipids Cat#850150 Dodecyl-b-D-maltoside (DDM) Carl Roth Cat#69227-93-6 n-Decyl b-maltoside (DM) Glycon Biochemical Cat#D322LA Triton X-100 Merck Cat#9036-19-5 Digitonin Sigma Aldrich Cat#D141 Tween 20 Roth Cat#P1379 Amylose resin NEB Cat#E8021L Pierce Glutathione Agarose Thermo Fisher Scientific Cat#16101 Pierce Detergent Removal Resin Thermo Fisher Scientific Cat#87780 Strep-Tactin Superflow Plus Qiagen Cat#30004 HisPur Cobalt Resin Thermo Fisher Scientific Cat#89964 Natriumhypodisulfit (Dithionite) Sigma Aldrich Cat#7775-14-6 Nycodenz Alere Technologies Cat#AXS-1002424 Slide-A-Lyzer Dialysekassetten, 10 K MWCO, 12 ml Thermo Fisher Scientific Cat#66807 SERVA FastLoad 1 kb DNA Ladder SERVA Cat#39317.01 SERVA Fastload 100 bp DNA Ladder SERVA Cat#39316.01 SERVA DNA Stain G SERVA Cat#39803 All Blue Prestained Protein Standards BioRad Cat#1610373 Protein Dual Color Standards BioRad Cat#1610374 Gel Loading Dye, Purple (6X) NEB Cat#B7024S BS3 (Bis(sulfosuccinimidyl)suberat) Thermo Fisher Scientific Cat#21580 Maltose Monohydrat Merck Cat#6363-53-7 L-Glutathion Merck Cat#70-18-8 X-TREMEGENE 9 DNA Tranfection Merck Cat#6365779001 Polycarbonate Membranes 0.4mm Avanti Cat#610007 Polycarbonate Membranes 0.1mm Avanti Cat#610005 10mm Filter Supports Avanti Cat#610014 Mini-PROTEAN TGX Stain-Free Protein Gels BioRad Cat#4568034 4–15% Mini-PROTEAN TGX Precast Protein Gels BioRad Cat#4561083 96-well microplate (non-binding) Greiner Bio-one Cat#655906 0.45um Syringe Filter AMSTAT Cat#C0000629 Amicon Ultra-15 concentrator 100 kDa Milipore Cat#UFC910024 Amicon Ultra-15 concentrator 30 kDa Milipore Cat#UFC903024 T5 exonuclease Epicentre Cat#T5E4111K BSA Roth Cat#8076 DAPI Roth Cat#6335 Phusion High-Fidelity DNA Polymerase NEB Cat#M0530 Q5 High-Fidelity DNA Polymerase NEB Cat#M0491 Taq DNA Polymerase NEB Cat#M0267 Taq DNA Ligase NEB Cat#M0208 BamHI NEB Cat#R0136S (Continued on next page) e2 Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023 .. REAGENT or RESOURCE SOURCE IDENTIFIER HindIII NEB Cat#R0104S Proteinase K Sigma Alrich Cat#39450-01-6 Benzonase Merck Cat#E1014 Dpn1 NEB Cat#R0176S Critical commercial assays QIAprep Spin Miniprep Kit Qiagen Cat#27106 Plasmid Mediprep Kit Invitrogen Cat#K210015 QIAquick PCR purification Kit Qiagen Cat#28106 HiPPR Detergent Removal Spin Column Kit Thermo Fisher Scientific Cat#PI88305 Atto 647N Protein Labeling Kit Jena Bioscience Cat#FP-201-647N Alexa Fluor 488 Protein Labeling Kit Invitrogen Cat#A10235 Zeba Spin-Entsalzungss€aulen, 7 K MWCO, 0,5 ml Thermo Fisher Scientific Cat#89882 LB medium BioChemica Cat#23143289 LB-agar Roth Cat#X969.2 ESF 921 medium Expression Systems Cat96-001-01 Sf-900 medium Gibco Cat#12658-019 Bafilomycin A1 Sigma-Aldrich Cat#B1793 DPBS Gibco Cat#14190-094 EBSS Gibco Cat#24010-043 GlutaMAX -I Gibco Cat#36600-021 FuGENE 6 Promega Cat#E2691 Polybrene Sigma Alrich Cat#H9268-106; Cat#sc-134220 puromycin InvivoGen Cat#ant-pr-1 blasticidin InvivoGen Cat#ant-bl-05 PhosSTOP Roche Cat#04906837001 Deposited data Uncropped Western blot and gels images, raw microscopy images This paper, Mendeley data https://doi.org/10.17632/dyy8xfvpkf.1 Experimental models: Cell lines Sf9 insect cells Invitrogen Cat#10503433 High Five insect cells Invitrogen Cat#10747474 Oligonucleotides Oligonucleotides used in this study This study Table S4 Recombinant DNA pLIB-6x-His-MBP-ATG9 This study N/A pLIB-ATG2-StrepII This study N/A pLIB-6xHis-GFP-WIPI4 This study N/A pLIB-GST-WIPI4 This study N/A pLIB-ATG101-StrepII This study N/A pBIG-6x-His-MBP-ATG13HORMA-ATG101 This study N/A pBIG-GST-ATG13HORMA-ATG101 This study N/A pLIB-6x-His-MBP-FIP200 This study N/A pLIB-6x-His-MBP-ATG13 This study N/A pLIB-6x-His-MBP-BECN1 This study N/A pLIB-GST-ULK1 This study N/A pLIB-6xHis-mCherry-ATG14 This study N/A pLIB-6x-His-MBP-ATG13HORMA This study N/A (Continued on next page) Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023 e3 .. REAGENT or RESOURCE SOURCE IDENTIFIER pLIB-GST- ATG13HORMA This study N/A pLIB-GST- ATG13HORMA_DSB1 This study N/A pLIB-6x-His-MBP- ATG13HORMA_DSB1 This study N/A pLIB-6x-His-MBP- ATG13HORMA_DSB2 This study N/A pLIB-6x-His-MBP-ATG101 This study N/A pLIB-GST-ATG101 This study N/A pLIB-GST-ATG9C This study N/A pLIB-GST-ATG9C, P833A, P834A This study N/A pLIB-6xHis-mCherry-ATG9N This study N/A pLIB-GST-ATG9N This study N/A pLIB-6x-His-MBP-ATG9N This study N/A pLIB-6xHis-GFP-ATG14 This study N/A pLIB-BECN1 This study N/A pLIB-6x-His-MBP-ATG9core This study N/A pLIB-6x-His-MBP-ATG9core+C This study N/A pLIB-6x-His-MBP-ATG9N+core This study N/A pBIG-GST-ATG13HORMA-6xHismCherry-ATG9N This study N/A pLIB-6x-His-MBP- ATG13HORMA_DN This study N/A pLIB-6x-His-MBP- ATG13LL This study N/A Software and algorithms JalView Barton Group (University of Dundee) 2.11.1.5 PyMol Schrödinger, LLC 2.5.2 AlphaFold 2 DeepMind (Alphabet Inc.) 2 Adobe Creative Cloud Adobe Inc. 5.8.0.592 Graphpad PRISM GraphPad Software Inc. 9.0.0 ImageJ NIH, USA 1.53t Image Studio Lite LI-COR Biosciences 5.2 ChimeraX 1.4 UCSF 1.4 Snapgene GSL Biotech 5.3.2 Fiji Schindelin et al.68 1.53 Microsoft Office Microsoft Corporation 16.64 Others MBPTrap HP (1 ml; 5 ml) Cytiva Cat#29048641; Cat#28918778 HiTrap TALON crude (1 ml; 5ml) Cytiva Cat#29048565; Cat#28953766 GSTrap HP (1 ml; 5 ml) Cytiva Cat#17528101; Cat#17528201 StrepTrap HP (5ml) Cytiva Cat#28907546 HiLoad 16/600 Superdex 75 pg Cytiva Cat#28989333 HiLoad 16/600 Superdex 200 pg Cytiva Cat#28989335 HiLoad 16/600 Superose 6 pg Cytiva Cat#29323952 Superdex 75 Increase 10/300 GL Cytiva Cat#29148721 Superdex 200 Increase 5/150 GL Cytiva Cat#28990945 Superose 6 Increase 10/300 GL Cytiva Cat#29091596 Superose 6 Increase 5/150 GL Cytiva Cat#29091597 RESOURCE Q, 6 ml Cytiva Cat#17117901 ProLong Glass Antifade Mountant Thermo Fisher Scientific Cat#P36980 e4 Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023
Purification:Article Title: Metamorphic proteins at the basis of human autophagy initiation and lipid transfer.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER 1,2-dioleoyl-sn-glycero-3phosphoethanolamine-N-(7-nitro-2-1,3benzoxadiazol-4-yl) (NBD-PE) Avanti Polar Lipids Cat#810145 1,2-dioleoyl-sn-glycero-3phosphoethanolamine-N-(lissamine rhodamine B sulfonyl) (RhoD-PE) Avanti Polar Lipids Cat#810150 1,2-dioleoyl-sn-glycero-3-phospho(1’-myo-inositol-3’-phosphate) (PI3P) Avanti Polar Lipids Cat#850150 Dodecyl-b-D-maltoside (DDM) Carl Roth Cat#69227-93-6 n-Decyl b-maltoside (DM) Glycon Biochemical Cat#D322LA Triton X-100 Merck Cat#9036-19-5 Digitonin Sigma Aldrich Cat#D141 Tween 20 Roth Cat#P1379 Amylose resin NEB Cat#E8021L Pierce Glutathione Agarose Thermo Fisher Scientific Cat#16101 Pierce Detergent Removal Resin Thermo Fisher Scientific Cat#87780 Strep-Tactin Superflow Plus Qiagen Cat#30004 HisPur Cobalt Resin Thermo Fisher Scientific Cat#89964 Natriumhypodisulfit (Dithionite) Sigma Aldrich Cat#7775-14-6 Nycodenz Alere Technologies Cat#AXS-1002424 Slide-A-Lyzer Dialysekassetten, 10 K MWCO, 12 ml Thermo Fisher Scientific Cat#66807 SERVA FastLoad 1 kb DNA Ladder SERVA Cat#39317.01 SERVA Fastload 100 bp DNA Ladder SERVA Cat#39316.01 SERVA DNA Stain G SERVA Cat#39803 All Blue Prestained Protein Standards BioRad Cat#1610373 Protein Dual Color Standards BioRad Cat#1610374 Gel Loading Dye, Purple (6X) NEB Cat#B7024S BS3 (Bis(sulfosuccinimidyl)suberat) Thermo Fisher Scientific Cat#21580 Maltose Monohydrat Merck Cat#6363-53-7 L-Glutathion Merck Cat#70-18-8 X-TREMEGENE 9 DNA Tranfection Merck Cat#6365779001 Polycarbonate Membranes 0.4mm Avanti Cat#610007 Polycarbonate Membranes 0.1mm Avanti Cat#610005 10mm Filter Supports Avanti Cat#610014 Mini-PROTEAN TGX Stain-Free Protein Gels BioRad Cat#4568034 4–15% Mini-PROTEAN TGX Precast Protein Gels BioRad Cat#4561083 96-well microplate (non-binding) Greiner Bio-one Cat#655906 0.45um Syringe Filter AMSTAT Cat#C0000629 Amicon Ultra-15 concentrator 100 kDa Milipore Cat#UFC910024 Amicon Ultra-15 concentrator 30 kDa Milipore Cat#UFC903024 T5 exonuclease Epicentre Cat#T5E4111K BSA Roth Cat#8076 DAPI Roth Cat#6335 Phusion High-Fidelity DNA Polymerase NEB Cat#M0530 Q5 High-Fidelity DNA Polymerase NEB Cat#M0491 Taq DNA Polymerase NEB Cat#M0267 Taq DNA Ligase NEB Cat#M0208 BamHI NEB Cat#R0136S (Continued on next page) e2 Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023 .. REAGENT or RESOURCE SOURCE IDENTIFIER HindIII NEB Cat#R0104S Proteinase K Sigma Alrich Cat#39450-01-6 Benzonase Merck Cat#E1014 Dpn1 NEB Cat#R0176S Critical commercial assays QIAprep Spin Miniprep Kit Qiagen Cat#27106 Plasmid Mediprep Kit Invitrogen Cat#K210015 QIAquick PCR purification Kit Qiagen Cat#28106 HiPPR Detergent Removal Spin Column Kit Thermo Fisher Scientific Cat#PI88305 Atto 647N Protein Labeling Kit Jena Bioscience Cat#FP-201-647N Alexa Fluor 488 Protein Labeling Kit Invitrogen Cat#A10235 Zeba Spin-Entsalzungss€aulen, 7 K MWCO, 0,5 ml Thermo Fisher Scientific Cat#89882 LB medium BioChemica Cat#23143289 LB-agar Roth Cat#X969.2 ESF 921 medium Expression Systems Cat96-001-01 Sf-900 medium Gibco Cat#12658-019 Bafilomycin A1 Sigma-Aldrich Cat#B1793 DPBS Gibco Cat#14190-094 EBSS Gibco Cat#24010-043 GlutaMAX -I Gibco Cat#36600-021 FuGENE 6 Promega Cat#E2691 Polybrene Sigma Alrich Cat#H9268-106; Cat#sc-134220 puromycin InvivoGen Cat#ant-pr-1 blasticidin InvivoGen Cat#ant-bl-05 PhosSTOP Roche Cat#04906837001 Deposited data Uncropped Western blot and gels images, raw microscopy images This paper, Mendeley data https://doi.org/10.17632/dyy8xfvpkf.1 Experimental models: Cell lines Sf9 insect cells Invitrogen Cat#10503433 High Five insect cells Invitrogen Cat#10747474 Oligonucleotides Oligonucleotides used in this study This study Table S4 Recombinant DNA pLIB-6x-His-MBP-ATG9 This study N/A pLIB-ATG2-StrepII This study N/A pLIB-6xHis-GFP-WIPI4 This study N/A pLIB-GST-WIPI4 This study N/A pLIB-ATG101-StrepII This study N/A pBIG-6x-His-MBP-ATG13HORMA-ATG101 This study N/A pBIG-GST-ATG13HORMA-ATG101 This study N/A pLIB-6x-His-MBP-FIP200 This study N/A pLIB-6x-His-MBP-ATG13 This study N/A pLIB-6x-His-MBP-BECN1 This study N/A pLIB-GST-ULK1 This study N/A pLIB-6xHis-mCherry-ATG14 This study N/A pLIB-6x-His-MBP-ATG13HORMA This study N/A (Continued on next page) Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023 e3 .. REAGENT or RESOURCE SOURCE IDENTIFIER pLIB-GST- ATG13HORMA This study N/A pLIB-GST- ATG13HORMA_DSB1 This study N/A pLIB-6x-His-MBP- ATG13HORMA_DSB1 This study N/A pLIB-6x-His-MBP- ATG13HORMA_DSB2 This study N/A pLIB-6x-His-MBP-ATG101 This study N/A pLIB-GST-ATG101 This study N/A pLIB-GST-ATG9C This study N/A pLIB-GST-ATG9C, P833A, P834A This study N/A pLIB-6xHis-mCherry-ATG9N This study N/A pLIB-GST-ATG9N This study N/A pLIB-6x-His-MBP-ATG9N This study N/A pLIB-6xHis-GFP-ATG14 This study N/A pLIB-BECN1 This study N/A pLIB-6x-His-MBP-ATG9core This study N/A pLIB-6x-His-MBP-ATG9core+C This study N/A pLIB-6x-His-MBP-ATG9N+core This study N/A pBIG-GST-ATG13HORMA-6xHismCherry-ATG9N This study N/A pLIB-6x-His-MBP- ATG13HORMA_DN This study N/A pLIB-6x-His-MBP- ATG13LL This study N/A Software and algorithms JalView Barton Group (University of Dundee) 2.11.1.5 PyMol Schrödinger, LLC 2.5.2 AlphaFold 2 DeepMind (Alphabet Inc.) 2 Adobe Creative Cloud Adobe Inc. 5.8.0.592 Graphpad PRISM GraphPad Software Inc. 9.0.0 ImageJ NIH, USA 1.53t Image Studio Lite LI-COR Biosciences 5.2 ChimeraX 1.4 UCSF 1.4 Snapgene GSL Biotech 5.3.2 Fiji Schindelin et al.68 1.53 Microsoft Office Microsoft Corporation 16.64 Others MBPTrap HP (1 ml; 5 ml) Cytiva Cat#29048641; Cat#28918778 HiTrap TALON crude (1 ml; 5ml) Cytiva Cat#29048565; Cat#28953766 GSTrap HP (1 ml; 5 ml) Cytiva Cat#17528101; Cat#17528201 StrepTrap HP (5ml) Cytiva Cat#28907546 HiLoad 16/600 Superdex 75 pg Cytiva Cat#28989333 HiLoad 16/600 Superdex 200 pg Cytiva Cat#28989335 HiLoad 16/600 Superose 6 pg Cytiva Cat#29323952 Superdex 75 Increase 10/300 GL Cytiva Cat#29148721 Superdex 200 Increase 5/150 GL Cytiva Cat#28990945 Superose 6 Increase 10/300 GL Cytiva Cat#29091596 Superose 6 Increase 5/150 GL Cytiva Cat#29091597 RESOURCE Q, 6 ml Cytiva Cat#17117901 ProLong Glass Antifade Mountant Thermo Fisher Scientific Cat#P36980 e4 Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023
Labeling:Article Title: Metamorphic proteins at the basis of human autophagy initiation and lipid transfer.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER 1,2-dioleoyl-sn-glycero-3phosphoethanolamine-N-(7-nitro-2-1,3benzoxadiazol-4-yl) (NBD-PE) Avanti Polar Lipids Cat#810145 1,2-dioleoyl-sn-glycero-3phosphoethanolamine-N-(lissamine rhodamine B sulfonyl) (RhoD-PE) Avanti Polar Lipids Cat#810150 1,2-dioleoyl-sn-glycero-3-phospho(1’-myo-inositol-3’-phosphate) (PI3P) Avanti Polar Lipids Cat#850150 Dodecyl-b-D-maltoside (DDM) Carl Roth Cat#69227-93-6 n-Decyl b-maltoside (DM) Glycon Biochemical Cat#D322LA Triton X-100 Merck Cat#9036-19-5 Digitonin Sigma Aldrich Cat#D141 Tween 20 Roth Cat#P1379 Amylose resin NEB Cat#E8021L Pierce Glutathione Agarose Thermo Fisher Scientific Cat#16101 Pierce Detergent Removal Resin Thermo Fisher Scientific Cat#87780 Strep-Tactin Superflow Plus Qiagen Cat#30004 HisPur Cobalt Resin Thermo Fisher Scientific Cat#89964 Natriumhypodisulfit (Dithionite) Sigma Aldrich Cat#7775-14-6 Nycodenz Alere Technologies Cat#AXS-1002424 Slide-A-Lyzer Dialysekassetten, 10 K MWCO, 12 ml Thermo Fisher Scientific Cat#66807 SERVA FastLoad 1 kb DNA Ladder SERVA Cat#39317.01 SERVA Fastload 100 bp DNA Ladder SERVA Cat#39316.01 SERVA DNA Stain G SERVA Cat#39803 All Blue Prestained Protein Standards BioRad Cat#1610373 Protein Dual Color Standards BioRad Cat#1610374 Gel Loading Dye, Purple (6X) NEB Cat#B7024S BS3 (Bis(sulfosuccinimidyl)suberat) Thermo Fisher Scientific Cat#21580 Maltose Monohydrat Merck Cat#6363-53-7 L-Glutathion Merck Cat#70-18-8 X-TREMEGENE 9 DNA Tranfection Merck Cat#6365779001 Polycarbonate Membranes 0.4mm Avanti Cat#610007 Polycarbonate Membranes 0.1mm Avanti Cat#610005 10mm Filter Supports Avanti Cat#610014 Mini-PROTEAN TGX Stain-Free Protein Gels BioRad Cat#4568034 4–15% Mini-PROTEAN TGX Precast Protein Gels BioRad Cat#4561083 96-well microplate (non-binding) Greiner Bio-one Cat#655906 0.45um Syringe Filter AMSTAT Cat#C0000629 Amicon Ultra-15 concentrator 100 kDa Milipore Cat#UFC910024 Amicon Ultra-15 concentrator 30 kDa Milipore Cat#UFC903024 T5 exonuclease Epicentre Cat#T5E4111K BSA Roth Cat#8076 DAPI Roth Cat#6335 Phusion High-Fidelity DNA Polymerase NEB Cat#M0530 Q5 High-Fidelity DNA Polymerase NEB Cat#M0491 Taq DNA Polymerase NEB Cat#M0267 Taq DNA Ligase NEB Cat#M0208 BamHI NEB Cat#R0136S (Continued on next page) e2 Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023 .. REAGENT or RESOURCE SOURCE IDENTIFIER HindIII NEB Cat#R0104S Proteinase K Sigma Alrich Cat#39450-01-6 Benzonase Merck Cat#E1014 Dpn1 NEB Cat#R0176S Critical commercial assays QIAprep Spin Miniprep Kit Qiagen Cat#27106 Plasmid Mediprep Kit Invitrogen Cat#K210015 QIAquick PCR purification Kit Qiagen Cat#28106 HiPPR Detergent Removal Spin Column Kit Thermo Fisher Scientific Cat#PI88305 Atto 647N Protein Labeling Kit Jena Bioscience Cat#FP-201-647N Alexa Fluor 488 Protein Labeling Kit Invitrogen Cat#A10235 Zeba Spin-Entsalzungss€aulen, 7 K MWCO, 0,5 ml Thermo Fisher Scientific Cat#89882 LB medium BioChemica Cat#23143289 LB-agar Roth Cat#X969.2 ESF 921 medium Expression Systems Cat96-001-01 Sf-900 medium Gibco Cat#12658-019 Bafilomycin A1 Sigma-Aldrich Cat#B1793 DPBS Gibco Cat#14190-094 EBSS Gibco Cat#24010-043 GlutaMAX -I Gibco Cat#36600-021 FuGENE 6 Promega Cat#E2691 Polybrene Sigma Alrich Cat#H9268-106; Cat#sc-134220 puromycin InvivoGen Cat#ant-pr-1 blasticidin InvivoGen Cat#ant-bl-05 PhosSTOP Roche Cat#04906837001 Deposited data Uncropped Western blot and gels images, raw microscopy images This paper, Mendeley data https://doi.org/10.17632/dyy8xfvpkf.1 Experimental models: Cell lines Sf9 insect cells Invitrogen Cat#10503433 High Five insect cells Invitrogen Cat#10747474 Oligonucleotides Oligonucleotides used in this study This study Table S4 Recombinant DNA pLIB-6x-His-MBP-ATG9 This study N/A pLIB-ATG2-StrepII This study N/A pLIB-6xHis-GFP-WIPI4 This study N/A pLIB-GST-WIPI4 This study N/A pLIB-ATG101-StrepII This study N/A pBIG-6x-His-MBP-ATG13HORMA-ATG101 This study N/A pBIG-GST-ATG13HORMA-ATG101 This study N/A pLIB-6x-His-MBP-FIP200 This study N/A pLIB-6x-His-MBP-ATG13 This study N/A pLIB-6x-His-MBP-BECN1 This study N/A pLIB-GST-ULK1 This study N/A pLIB-6xHis-mCherry-ATG14 This study N/A pLIB-6x-His-MBP-ATG13HORMA This study N/A (Continued on next page) Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023 e3 .. REAGENT or RESOURCE SOURCE IDENTIFIER pLIB-GST- ATG13HORMA This study N/A pLIB-GST- ATG13HORMA_DSB1 This study N/A pLIB-6x-His-MBP- ATG13HORMA_DSB1 This study N/A pLIB-6x-His-MBP- ATG13HORMA_DSB2 This study N/A pLIB-6x-His-MBP-ATG101 This study N/A pLIB-GST-ATG101 This study N/A pLIB-GST-ATG9C This study N/A pLIB-GST-ATG9C, P833A, P834A This study N/A pLIB-6xHis-mCherry-ATG9N This study N/A pLIB-GST-ATG9N This study N/A pLIB-6x-His-MBP-ATG9N This study N/A pLIB-6xHis-GFP-ATG14 This study N/A pLIB-BECN1 This study N/A pLIB-6x-His-MBP-ATG9core This study N/A pLIB-6x-His-MBP-ATG9core+C This study N/A pLIB-6x-His-MBP-ATG9N+core This study N/A pBIG-GST-ATG13HORMA-6xHismCherry-ATG9N This study N/A pLIB-6x-His-MBP- ATG13HORMA_DN This study N/A pLIB-6x-His-MBP- ATG13LL This study N/A Software and algorithms JalView Barton Group (University of Dundee) 2.11.1.5 PyMol Schrödinger, LLC 2.5.2 AlphaFold 2 DeepMind (Alphabet Inc.) 2 Adobe Creative Cloud Adobe Inc. 5.8.0.592 Graphpad PRISM GraphPad Software Inc. 9.0.0 ImageJ NIH, USA 1.53t Image Studio Lite LI-COR Biosciences 5.2 ChimeraX 1.4 UCSF 1.4 Snapgene GSL Biotech 5.3.2 Fiji Schindelin et al.68 1.53 Microsoft Office Microsoft Corporation 16.64 Others MBPTrap HP (1 ml; 5 ml) Cytiva Cat#29048641; Cat#28918778 HiTrap TALON crude (1 ml; 5ml) Cytiva Cat#29048565; Cat#28953766 GSTrap HP (1 ml; 5 ml) Cytiva Cat#17528101; Cat#17528201 StrepTrap HP (5ml) Cytiva Cat#28907546 HiLoad 16/600 Superdex 75 pg Cytiva Cat#28989333 HiLoad 16/600 Superdex 200 pg Cytiva Cat#28989335 HiLoad 16/600 Superose 6 pg Cytiva Cat#29323952 Superdex 75 Increase 10/300 GL Cytiva Cat#29148721 Superdex 200 Increase 5/150 GL Cytiva Cat#28990945 Superose 6 Increase 10/300 GL Cytiva Cat#29091596 Superose 6 Increase 5/150 GL Cytiva Cat#29091597 RESOURCE Q, 6 ml Cytiva Cat#17117901 ProLong Glass Antifade Mountant Thermo Fisher Scientific Cat#P36980 e4 Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023
Western Blot:Article Title: Metamorphic proteins at the basis of human autophagy initiation and lipid transfer.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER 1,2-dioleoyl-sn-glycero-3phosphoethanolamine-N-(7-nitro-2-1,3benzoxadiazol-4-yl) (NBD-PE) Avanti Polar Lipids Cat#810145 1,2-dioleoyl-sn-glycero-3phosphoethanolamine-N-(lissamine rhodamine B sulfonyl) (RhoD-PE) Avanti Polar Lipids Cat#810150 1,2-dioleoyl-sn-glycero-3-phospho(1’-myo-inositol-3’-phosphate) (PI3P) Avanti Polar Lipids Cat#850150 Dodecyl-b-D-maltoside (DDM) Carl Roth Cat#69227-93-6 n-Decyl b-maltoside (DM) Glycon Biochemical Cat#D322LA Triton X-100 Merck Cat#9036-19-5 Digitonin Sigma Aldrich Cat#D141 Tween 20 Roth Cat#P1379 Amylose resin NEB Cat#E8021L Pierce Glutathione Agarose Thermo Fisher Scientific Cat#16101 Pierce Detergent Removal Resin Thermo Fisher Scientific Cat#87780 Strep-Tactin Superflow Plus Qiagen Cat#30004 HisPur Cobalt Resin Thermo Fisher Scientific Cat#89964 Natriumhypodisulfit (Dithionite) Sigma Aldrich Cat#7775-14-6 Nycodenz Alere Technologies Cat#AXS-1002424 Slide-A-Lyzer Dialysekassetten, 10 K MWCO, 12 ml Thermo Fisher Scientific Cat#66807 SERVA FastLoad 1 kb DNA Ladder SERVA Cat#39317.01 SERVA Fastload 100 bp DNA Ladder SERVA Cat#39316.01 SERVA DNA Stain G SERVA Cat#39803 All Blue Prestained Protein Standards BioRad Cat#1610373 Protein Dual Color Standards BioRad Cat#1610374 Gel Loading Dye, Purple (6X) NEB Cat#B7024S BS3 (Bis(sulfosuccinimidyl)suberat) Thermo Fisher Scientific Cat#21580 Maltose Monohydrat Merck Cat#6363-53-7 L-Glutathion Merck Cat#70-18-8 X-TREMEGENE 9 DNA Tranfection Merck Cat#6365779001 Polycarbonate Membranes 0.4mm Avanti Cat#610007 Polycarbonate Membranes 0.1mm Avanti Cat#610005 10mm Filter Supports Avanti Cat#610014 Mini-PROTEAN TGX Stain-Free Protein Gels BioRad Cat#4568034 4–15% Mini-PROTEAN TGX Precast Protein Gels BioRad Cat#4561083 96-well microplate (non-binding) Greiner Bio-one Cat#655906 0.45um Syringe Filter AMSTAT Cat#C0000629 Amicon Ultra-15 concentrator 100 kDa Milipore Cat#UFC910024 Amicon Ultra-15 concentrator 30 kDa Milipore Cat#UFC903024 T5 exonuclease Epicentre Cat#T5E4111K BSA Roth Cat#8076 DAPI Roth Cat#6335 Phusion High-Fidelity DNA Polymerase NEB Cat#M0530 Q5 High-Fidelity DNA Polymerase NEB Cat#M0491 Taq DNA Polymerase NEB Cat#M0267 Taq DNA Ligase NEB Cat#M0208 BamHI NEB Cat#R0136S (Continued on next page) e2 Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023 .. REAGENT or RESOURCE SOURCE IDENTIFIER HindIII NEB Cat#R0104S Proteinase K Sigma Alrich Cat#39450-01-6 Benzonase Merck Cat#E1014 Dpn1 NEB Cat#R0176S Critical commercial assays QIAprep Spin Miniprep Kit Qiagen Cat#27106 Plasmid Mediprep Kit Invitrogen Cat#K210015 QIAquick PCR purification Kit Qiagen Cat#28106 HiPPR Detergent Removal Spin Column Kit Thermo Fisher Scientific Cat#PI88305 Atto 647N Protein Labeling Kit Jena Bioscience Cat#FP-201-647N Alexa Fluor 488 Protein Labeling Kit Invitrogen Cat#A10235 Zeba Spin-Entsalzungss€aulen, 7 K MWCO, 0,5 ml Thermo Fisher Scientific Cat#89882 LB medium BioChemica Cat#23143289 LB-agar Roth Cat#X969.2 ESF 921 medium Expression Systems Cat96-001-01 Sf-900 medium Gibco Cat#12658-019 Bafilomycin A1 Sigma-Aldrich Cat#B1793 DPBS Gibco Cat#14190-094 EBSS Gibco Cat#24010-043 GlutaMAX -I Gibco Cat#36600-021 FuGENE 6 Promega Cat#E2691 Polybrene Sigma Alrich Cat#H9268-106; Cat#sc-134220 puromycin InvivoGen Cat#ant-pr-1 blasticidin InvivoGen Cat#ant-bl-05 PhosSTOP Roche Cat#04906837001 Deposited data Uncropped Western blot and gels images, raw microscopy images This paper, Mendeley data https://doi.org/10.17632/dyy8xfvpkf.1 Experimental models: Cell lines Sf9 insect cells Invitrogen Cat#10503433 High Five insect cells Invitrogen Cat#10747474 Oligonucleotides Oligonucleotides used in this study This study Table S4 Recombinant DNA pLIB-6x-His-MBP-ATG9 This study N/A pLIB-ATG2-StrepII This study N/A pLIB-6xHis-GFP-WIPI4 This study N/A pLIB-GST-WIPI4 This study N/A pLIB-ATG101-StrepII This study N/A pBIG-6x-His-MBP-ATG13HORMA-ATG101 This study N/A pBIG-GST-ATG13HORMA-ATG101 This study N/A pLIB-6x-His-MBP-FIP200 This study N/A pLIB-6x-His-MBP-ATG13 This study N/A pLIB-6x-His-MBP-BECN1 This study N/A pLIB-GST-ULK1 This study N/A pLIB-6xHis-mCherry-ATG14 This study N/A pLIB-6x-His-MBP-ATG13HORMA This study N/A (Continued on next page) Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023 e3 .. REAGENT or RESOURCE SOURCE IDENTIFIER pLIB-GST- ATG13HORMA This study N/A pLIB-GST- ATG13HORMA_DSB1 This study N/A pLIB-6x-His-MBP- ATG13HORMA_DSB1 This study N/A pLIB-6x-His-MBP- ATG13HORMA_DSB2 This study N/A pLIB-6x-His-MBP-ATG101 This study N/A pLIB-GST-ATG101 This study N/A pLIB-GST-ATG9C This study N/A pLIB-GST-ATG9C, P833A, P834A This study N/A pLIB-6xHis-mCherry-ATG9N This study N/A pLIB-GST-ATG9N This study N/A pLIB-6x-His-MBP-ATG9N This study N/A pLIB-6xHis-GFP-ATG14 This study N/A pLIB-BECN1 This study N/A pLIB-6x-His-MBP-ATG9core This study N/A pLIB-6x-His-MBP-ATG9core+C This study N/A pLIB-6x-His-MBP-ATG9N+core This study N/A pBIG-GST-ATG13HORMA-6xHismCherry-ATG9N This study N/A pLIB-6x-His-MBP- ATG13HORMA_DN This study N/A pLIB-6x-His-MBP- ATG13LL This study N/A Software and algorithms JalView Barton Group (University of Dundee) 2.11.1.5 PyMol Schrödinger, LLC 2.5.2 AlphaFold 2 DeepMind (Alphabet Inc.) 2 Adobe Creative Cloud Adobe Inc. 5.8.0.592 Graphpad PRISM GraphPad Software Inc. 9.0.0 ImageJ NIH, USA 1.53t Image Studio Lite LI-COR Biosciences 5.2 ChimeraX 1.4 UCSF 1.4 Snapgene GSL Biotech 5.3.2 Fiji Schindelin et al.68 1.53 Microsoft Office Microsoft Corporation 16.64 Others MBPTrap HP (1 ml; 5 ml) Cytiva Cat#29048641; Cat#28918778 HiTrap TALON crude (1 ml; 5ml) Cytiva Cat#29048565; Cat#28953766 GSTrap HP (1 ml; 5 ml) Cytiva Cat#17528101; Cat#17528201 StrepTrap HP (5ml) Cytiva Cat#28907546 HiLoad 16/600 Superdex 75 pg Cytiva Cat#28989333 HiLoad 16/600 Superdex 200 pg Cytiva Cat#28989335 HiLoad 16/600 Superose 6 pg Cytiva Cat#29323952 Superdex 75 Increase 10/300 GL Cytiva Cat#29148721 Superdex 200 Increase 5/150 GL Cytiva Cat#28990945 Superose 6 Increase 10/300 GL Cytiva Cat#29091596 Superose 6 Increase 5/150 GL Cytiva Cat#29091597 RESOURCE Q, 6 ml Cytiva Cat#17117901 ProLong Glass Antifade Mountant Thermo Fisher Scientific Cat#P36980 e4 Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023
Microscopy:Article Title: Metamorphic proteins at the basis of human autophagy initiation and lipid transfer.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER 1,2-dioleoyl-sn-glycero-3phosphoethanolamine-N-(7-nitro-2-1,3benzoxadiazol-4-yl) (NBD-PE) Avanti Polar Lipids Cat#810145 1,2-dioleoyl-sn-glycero-3phosphoethanolamine-N-(lissamine rhodamine B sulfonyl) (RhoD-PE) Avanti Polar Lipids Cat#810150 1,2-dioleoyl-sn-glycero-3-phospho(1’-myo-inositol-3’-phosphate) (PI3P) Avanti Polar Lipids Cat#850150 Dodecyl-b-D-maltoside (DDM) Carl Roth Cat#69227-93-6 n-Decyl b-maltoside (DM) Glycon Biochemical Cat#D322LA Triton X-100 Merck Cat#9036-19-5 Digitonin Sigma Aldrich Cat#D141 Tween 20 Roth Cat#P1379 Amylose resin NEB Cat#E8021L Pierce Glutathione Agarose Thermo Fisher Scientific Cat#16101 Pierce Detergent Removal Resin Thermo Fisher Scientific Cat#87780 Strep-Tactin Superflow Plus Qiagen Cat#30004 HisPur Cobalt Resin Thermo Fisher Scientific Cat#89964 Natriumhypodisulfit (Dithionite) Sigma Aldrich Cat#7775-14-6 Nycodenz Alere Technologies Cat#AXS-1002424 Slide-A-Lyzer Dialysekassetten, 10 K MWCO, 12 ml Thermo Fisher Scientific Cat#66807 SERVA FastLoad 1 kb DNA Ladder SERVA Cat#39317.01 SERVA Fastload 100 bp DNA Ladder SERVA Cat#39316.01 SERVA DNA Stain G SERVA Cat#39803 All Blue Prestained Protein Standards BioRad Cat#1610373 Protein Dual Color Standards BioRad Cat#1610374 Gel Loading Dye, Purple (6X) NEB Cat#B7024S BS3 (Bis(sulfosuccinimidyl)suberat) Thermo Fisher Scientific Cat#21580 Maltose Monohydrat Merck Cat#6363-53-7 L-Glutathion Merck Cat#70-18-8 X-TREMEGENE 9 DNA Tranfection Merck Cat#6365779001 Polycarbonate Membranes 0.4mm Avanti Cat#610007 Polycarbonate Membranes 0.1mm Avanti Cat#610005 10mm Filter Supports Avanti Cat#610014 Mini-PROTEAN TGX Stain-Free Protein Gels BioRad Cat#4568034 4–15% Mini-PROTEAN TGX Precast Protein Gels BioRad Cat#4561083 96-well microplate (non-binding) Greiner Bio-one Cat#655906 0.45um Syringe Filter AMSTAT Cat#C0000629 Amicon Ultra-15 concentrator 100 kDa Milipore Cat#UFC910024 Amicon Ultra-15 concentrator 30 kDa Milipore Cat#UFC903024 T5 exonuclease Epicentre Cat#T5E4111K BSA Roth Cat#8076 DAPI Roth Cat#6335 Phusion High-Fidelity DNA Polymerase NEB Cat#M0530 Q5 High-Fidelity DNA Polymerase NEB Cat#M0491 Taq DNA Polymerase NEB Cat#M0267 Taq DNA Ligase NEB Cat#M0208 BamHI NEB Cat#R0136S (Continued on next page) e2 Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023 .. REAGENT or RESOURCE SOURCE IDENTIFIER HindIII NEB Cat#R0104S Proteinase K Sigma Alrich Cat#39450-01-6 Benzonase Merck Cat#E1014 Dpn1 NEB Cat#R0176S Critical commercial assays QIAprep Spin Miniprep Kit Qiagen Cat#27106 Plasmid Mediprep Kit Invitrogen Cat#K210015 QIAquick PCR purification Kit Qiagen Cat#28106 HiPPR Detergent Removal Spin Column Kit Thermo Fisher Scientific Cat#PI88305 Atto 647N Protein Labeling Kit Jena Bioscience Cat#FP-201-647N Alexa Fluor 488 Protein Labeling Kit Invitrogen Cat#A10235 Zeba Spin-Entsalzungss€aulen, 7 K MWCO, 0,5 ml Thermo Fisher Scientific Cat#89882 LB medium BioChemica Cat#23143289 LB-agar Roth Cat#X969.2 ESF 921 medium Expression Systems Cat96-001-01 Sf-900 medium Gibco Cat#12658-019 Bafilomycin A1 Sigma-Aldrich Cat#B1793 DPBS Gibco Cat#14190-094 EBSS Gibco Cat#24010-043 GlutaMAX -I Gibco Cat#36600-021 FuGENE 6 Promega Cat#E2691 Polybrene Sigma Alrich Cat#H9268-106; Cat#sc-134220 puromycin InvivoGen Cat#ant-pr-1 blasticidin InvivoGen Cat#ant-bl-05 PhosSTOP Roche Cat#04906837001 Deposited data Uncropped Western blot and gels images, raw microscopy images This paper, Mendeley data https://doi.org/10.17632/dyy8xfvpkf.1 Experimental models: Cell lines Sf9 insect cells Invitrogen Cat#10503433 High Five insect cells Invitrogen Cat#10747474 Oligonucleotides Oligonucleotides used in this study This study Table S4 Recombinant DNA pLIB-6x-His-MBP-ATG9 This study N/A pLIB-ATG2-StrepII This study N/A pLIB-6xHis-GFP-WIPI4 This study N/A pLIB-GST-WIPI4 This study N/A pLIB-ATG101-StrepII This study N/A pBIG-6x-His-MBP-ATG13HORMA-ATG101 This study N/A pBIG-GST-ATG13HORMA-ATG101 This study N/A pLIB-6x-His-MBP-FIP200 This study N/A pLIB-6x-His-MBP-ATG13 This study N/A pLIB-6x-His-MBP-BECN1 This study N/A pLIB-GST-ULK1 This study N/A pLIB-6xHis-mCherry-ATG14 This study N/A pLIB-6x-His-MBP-ATG13HORMA This study N/A (Continued on next page) Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023 e3 .. REAGENT or RESOURCE SOURCE IDENTIFIER pLIB-GST- ATG13HORMA This study N/A pLIB-GST- ATG13HORMA_DSB1 This study N/A pLIB-6x-His-MBP- ATG13HORMA_DSB1 This study N/A pLIB-6x-His-MBP- ATG13HORMA_DSB2 This study N/A pLIB-6x-His-MBP-ATG101 This study N/A pLIB-GST-ATG101 This study N/A pLIB-GST-ATG9C This study N/A pLIB-GST-ATG9C, P833A, P834A This study N/A pLIB-6xHis-mCherry-ATG9N This study N/A pLIB-GST-ATG9N This study N/A pLIB-6x-His-MBP-ATG9N This study N/A pLIB-6xHis-GFP-ATG14 This study N/A pLIB-BECN1 This study N/A pLIB-6x-His-MBP-ATG9core This study N/A pLIB-6x-His-MBP-ATG9core+C This study N/A pLIB-6x-His-MBP-ATG9N+core This study N/A pBIG-GST-ATG13HORMA-6xHismCherry-ATG9N This study N/A pLIB-6x-His-MBP- ATG13HORMA_DN This study N/A pLIB-6x-His-MBP- ATG13LL This study N/A Software and algorithms JalView Barton Group (University of Dundee) 2.11.1.5 PyMol Schrödinger, LLC 2.5.2 AlphaFold 2 DeepMind (Alphabet Inc.) 2 Adobe Creative Cloud Adobe Inc. 5.8.0.592 Graphpad PRISM GraphPad Software Inc. 9.0.0 ImageJ NIH, USA 1.53t Image Studio Lite LI-COR Biosciences 5.2 ChimeraX 1.4 UCSF 1.4 Snapgene GSL Biotech 5.3.2 Fiji Schindelin et al.68 1.53 Microsoft Office Microsoft Corporation 16.64 Others MBPTrap HP (1 ml; 5 ml) Cytiva Cat#29048641; Cat#28918778 HiTrap TALON crude (1 ml; 5ml) Cytiva Cat#29048565; Cat#28953766 GSTrap HP (1 ml; 5 ml) Cytiva Cat#17528101; Cat#17528201 StrepTrap HP (5ml) Cytiva Cat#28907546 HiLoad 16/600 Superdex 75 pg Cytiva Cat#28989333 HiLoad 16/600 Superdex 200 pg Cytiva Cat#28989335 HiLoad 16/600 Superose 6 pg Cytiva Cat#29323952 Superdex 75 Increase 10/300 GL Cytiva Cat#29148721 Superdex 200 Increase 5/150 GL Cytiva Cat#28990945 Superose 6 Increase 10/300 GL Cytiva Cat#29091596 Superose 6 Increase 5/150 GL Cytiva Cat#29091597 RESOURCE Q, 6 ml Cytiva Cat#17117901 ProLong Glass Antifade Mountant Thermo Fisher Scientific Cat#P36980 e4 Molecular Cell 83, 2077–2090.e1–e12, June 15, 2023
Expressing:Article Title: Activation and closed-state inactivation mechanisms of the human voltage-gated K V 4 channel complexes.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Bacterial strains Turbo competent cells New England Biolabs Cat#C2984H DH10Bac competent cells Thermo Fisher Scientific Cat#10361012 Chemicals, peptides, and recombinant proteins Protease inhibitor cocktail MedChemExpress Cat#HY-K0010 Lauryl maltose neopentyl glycol (LMNG) Anatrace Cat#NG310 n-Dodecyl-b-D-Maltopyranoside (DDM) Anatrace Cat#D310S Glyco-Diosgenin (GDN) Anatrace Cat#GDN101 Cholesteryl hemisuccinate tris salt Anatrace Cat#CH210 ESF 921 Medium Expression Systems Cat96-001-01 Freestyle 293 expression medium Gibco Cat#12338018 DMEM/F-12 medium Gibco Cat#11330-032 Penicillin-streptomycin solution, 100x Corning Cat#30-002-CI Fetal bovine serum Gibco Cat#16140089 Lipofectamine 2000 reagent Invitrogen Cat#11668019 TransIT-Insect transfection reagent Mirus Cat#MIR6100 1,1-Methanediyl Bismethanethiosulfonate (MTS-1-MTS) Santa Cruz Biotech Cat#SC-208706 1,4-Butanediyl Bismethanethiosulfonate (MTS-4-MTS) Santa Cruz Biotech Cat#SC-208789 Deposited data KV4.2/KChIP2 channel complex, open state This study PDB: 7UK5; EMDB: EMD-26575 KV4.2/KChIP2 channel complex, inactivated state, class 1 This study PDB: 7UKC; EMDB: EMD-26576 KV4.2/KChIP2 channel complex, inactivated state, class 2 This study PDB: 7UKD; EMDB: EMD-26577 KV4.2/KChIP2 channel complex, intermediate state This study PDB: 7UKE; EMDB: EMD-26578 KV4.2/KChIP2 channel complex, putative resting state This study PDB: 7UKF; EMDB: EMD-26579 KV4.2/KChIP2/DPP6 channel complex, open state, transmembrane region This study PDB: 7UKG; EMDB: EMD-26580 KV4.2/KChIP2/DPP6 channel complex, open state, intracellular region This study PDB: 7UKH; EMDB: EMD-26581 Experimental models: Cell lines HEK293 ATCC Cat#CRL-1573 Sf9 ATCC Cat#CRL-1711 HEK293S GnTI– ATCC Cat#CRL-3022 Recombinant DNA pEG-BacMam Human Kv4.2EM This study N/A pEG-BacMam Human KChIP2 This study N/A pEG-BacMam Human DPP6 This study N/A Software and algorithms Serial EM Mastronarde (2005) http://bio3d.colorado.edu/SerialEM RELION Zivanov et al. (2018) https://github.com/3dem/relion (Continued on next page) Molecular Cell 82, 2427–2442.e1–e4, July 7, 2022 e1 .. REAGENT or RESOURCE SOURCE IDENTIFIER cryoSPARC Punjani et al. (2017) https://cryosparc.com/ Coot Emsley et al. (2010) http://www2.mrc-lmb.cam.ac.uk/personal/ pemsley/coot Phenix Afonine et al. (2018) https://www.phenix-online.org MolProbity Chen et al. (2010) http://molprobity.biochem.duke.edu Chimera Pettersen et al. (2004) https://www.cgl.ucsf.edu/chimera PyMOL Schrodinger http://www.pymol.org pCLAMP Molecular Devices https://www.moleculardevices.com/ Anaconda Anaconda https://www.anaconda.com/ Other CNBR-activated Sepharose beads Cytiva Cat#17043001 Superose 6 Increase 10/300 GL SEC column Cytiva Cat#29091596 R1.2/1.3 400 mesh UltrAuFoil gold grids Quantifoil Cat#Q350AR13A
Transfection:Article Title: Activation and closed-state inactivation mechanisms of the human voltage-gated K V 4 channel complexes.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Bacterial strains Turbo competent cells New England Biolabs Cat#C2984H DH10Bac competent cells Thermo Fisher Scientific Cat#10361012 Chemicals, peptides, and recombinant proteins Protease inhibitor cocktail MedChemExpress Cat#HY-K0010 Lauryl maltose neopentyl glycol (LMNG) Anatrace Cat#NG310 n-Dodecyl-b-D-Maltopyranoside (DDM) Anatrace Cat#D310S Glyco-Diosgenin (GDN) Anatrace Cat#GDN101 Cholesteryl hemisuccinate tris salt Anatrace Cat#CH210 ESF 921 Medium Expression Systems Cat96-001-01 Freestyle 293 expression medium Gibco Cat#12338018 DMEM/F-12 medium Gibco Cat#11330-032 Penicillin-streptomycin solution, 100x Corning Cat#30-002-CI Fetal bovine serum Gibco Cat#16140089 Lipofectamine 2000 reagent Invitrogen Cat#11668019 TransIT-Insect transfection reagent Mirus Cat#MIR6100 1,1-Methanediyl Bismethanethiosulfonate (MTS-1-MTS) Santa Cruz Biotech Cat#SC-208706 1,4-Butanediyl Bismethanethiosulfonate (MTS-4-MTS) Santa Cruz Biotech Cat#SC-208789 Deposited data KV4.2/KChIP2 channel complex, open state This study PDB: 7UK5; EMDB: EMD-26575 KV4.2/KChIP2 channel complex, inactivated state, class 1 This study PDB: 7UKC; EMDB: EMD-26576 KV4.2/KChIP2 channel complex, inactivated state, class 2 This study PDB: 7UKD; EMDB: EMD-26577 KV4.2/KChIP2 channel complex, intermediate state This study PDB: 7UKE; EMDB: EMD-26578 KV4.2/KChIP2 channel complex, putative resting state This study PDB: 7UKF; EMDB: EMD-26579 KV4.2/KChIP2/DPP6 channel complex, open state, transmembrane region This study PDB: 7UKG; EMDB: EMD-26580 KV4.2/KChIP2/DPP6 channel complex, open state, intracellular region This study PDB: 7UKH; EMDB: EMD-26581 Experimental models: Cell lines HEK293 ATCC Cat#CRL-1573 Sf9 ATCC Cat#CRL-1711 HEK293S GnTI– ATCC Cat#CRL-3022 Recombinant DNA pEG-BacMam Human Kv4.2EM This study N/A pEG-BacMam Human KChIP2 This study N/A pEG-BacMam Human DPP6 This study N/A Software and algorithms Serial EM Mastronarde (2005) http://bio3d.colorado.edu/SerialEM RELION Zivanov et al. (2018) https://github.com/3dem/relion (Continued on next page) Molecular Cell 82, 2427–2442.e1–e4, July 7, 2022 e1 .. REAGENT or RESOURCE SOURCE IDENTIFIER cryoSPARC Punjani et al. (2017) https://cryosparc.com/ Coot Emsley et al. (2010) http://www2.mrc-lmb.cam.ac.uk/personal/ pemsley/coot Phenix Afonine et al. (2018) https://www.phenix-online.org MolProbity Chen et al. (2010) http://molprobity.biochem.duke.edu Chimera Pettersen et al. (2004) https://www.cgl.ucsf.edu/chimera PyMOL Schrodinger http://www.pymol.org pCLAMP Molecular Devices https://www.moleculardevices.com/ Anaconda Anaconda https://www.anaconda.com/ Other CNBR-activated Sepharose beads Cytiva Cat#17043001 Superose 6 Increase 10/300 GL SEC column Cytiva Cat#29091596 R1.2/1.3 400 mesh UltrAuFoil gold grids Quantifoil Cat#Q350AR13A
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