9nc1

MicroED structure of papain-E-64 complex from microcrystals soaked with protease inhibitor cocktail

spinqualitylabelsall models 9nc1, resolution 2.40Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image   Print3D Structural highlights 9nc1 is a 1 chain structure with sequence from Carica papaya. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance. Method: Electron crystallography, Resolution 2.4Å Ligands: E64 Resources: FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT Function PAPA1_CARPA Publication Abstract from PubMed With the goal of accelerating the discovery of small molecule-protein complexes, we leverage fast, low-dose, event based electron counting microcrystal electron diffraction (MicroED) data collection and native mass spectrometry. This approach resolves structures of the epoxide-based cysteine protease inhibitor, and natural product, E-64, and its biosynthetic analogs bound to the model cysteine protease, papain. The combined structural power of MicroED and the analytical capabilities of native mass spectrometry (ED-MS) allows assignment of papain structures bound to E-64-like ligands with data obtained from crystal slurries soaked with mixtures of known inhibitors, and crude biosynthetic reactions. ED-MS further discriminates the highest-affinity ligand soaked into microcrystals from a broad inhibitor cocktail, and identifies multiple similarly high-affinity ligands soaked into microcrystals simultaneously. This extends to libraries of printed ligands dispensed directly onto TEM grids and later soaked with papain microcrystal slurries. ED-MS identifies papain binding to its preferred natural products, by showing that two analogues of E-64 outcompete others in binding to papain crystals, and by detecting papain bound to E-64 and an analogue from crude biosynthetic reactions, without purification. This illustrates the utility of ED-MS for natural product ligand discovery and for structure-based screening of small molecule binders to macromolecular targets. Combining MicroED and native mass spectrometry for structural discovery of enzyme-biosynthetic inhibitor complexes.,Vlahakis NW, Flowers CW, Liu M, Agdanowski M, Johnson S, Summers JA, Keyser C, Russell P, Rose S, Orlans J, Adhami N, Chen Y, Sawaya MR, Basu S, de Sanctis D, Wakatsuki S, Nelson HM, Loo JA, Tang Y, Rodriguez JA bioRxiv [Preprint]. 2025 Feb 26:2025.02.20.638743. doi: , 10.1101/2025.02.20.638743. PMID:40060639 From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.

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