1se9
Structure of At3g01050, a ubiquitin-fold protein from Arabidopsis thalianaStructure of At3g01050, a ubiquitin-fold protein from Arabidopsis thaliana
Structural highlights
FunctionMUB1_ARATH May serve as docking site to facilitate the association of other proteins to the plasma membrane. Evolutionary Conservation Check, as determined by ConSurfDB. You may read the explanation of the method and the full data available from ConSurf. Publication Abstract from PubMedStructural proteomics requires robust, scalable methods. Here we describe a wheat germ cell-free platform for protein production that supports efficient NMR structural studies of eukaryotic proteins and offers advantages over cell-based methods. To illustrate this platform, we describe its application to a specific target (At3g01050.1) from Arabidopsis thaliana. After cloning the target gene into a specialized plasmid, we carry out a small-scale (50 mul) in vitro sequential transcription and translation trial to ascertain the level of protein production and solubility. Next, we prepare mRNA for use in a 4-ml semicontinuous cell-free translation reaction to incorporate (15)N-labeled amino acids into a protein sample that we purify and test for suitability for NMR structural analysis. We then repeat the cell-free approach with (13)C,(15)N-labeled amino acids to prepare a doubly labeled sample. The three-dimensional (3D) structure of At3g01050.1 shows that this protein is an unusual member of the beta-grasp protein family. Cell-free protein production and labeling protocol for NMR-based structural proteomics.,Vinarov DA, Lytle BL, Peterson FC, Tyler EM, Volkman BF, Markley JL Nat Methods. 2004 Nov;1(2):149-53. Epub 2004 Oct 21. PMID:15782178[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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