1r50

Bacillus subtilis lipase A with covalently bound Sc-IPG-phosphonate-inhibitor

spinqualitylabelsall models 1r50, resolution 1.45Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image   Print3D Structural highlights 1r50 is a 2 chain structure with sequence from Bacillus subtilis. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance. Method: X-ray diffraction, Resolution 1.45Å Ligands: SIL Resources: FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT Function ESTA_BACSU Active toward p-nitrophenyl esters and triacylglycerides with a marked preference for esters with C8 acyl groups. 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 PubMed Phage display can be used as a protein-engineering tool for the selection of proteins with desirable binding properties from a library of mutants. Here we describe the application of this method for the directed evolution of Bacillus subtilis lipase A, an enzyme that has important properties for the preparation of the pharmaceutically relevant chiral compound 1,2-O-isopropylidene-sn-glycerol (IPG). PCR mutagenesis with spiked oligonucleotides was employed for saturation mutagenesis of a stretch of amino acids near the active site. After expression of these mutants on bacteriophages, dual selection with (S)-(+)- and (R)-(-)-IPG stereoisomers covalently coupled to enantiomeric phosphonate suicide inhibitors (SIRAN Sc and Rc inhibitors, respectively) was used for the isolation of variants with inverted enantioselectivity. The mutants were further characterised by determination of their Michaelis-Menten parameters. The 3D structures of the Sc and Rc inhibitor-lipase complexes were determined and provided structural insight into the mechanism of enantioselectivity of the enzyme. In conclusion, we have used phage display as a fast and reproducible method for the selection of Bacillus lipase A mutant enzymes with inverted enantioselectivity. Directed evolution of Bacillus subtilis lipase A by use of enantiomeric phosphonate inhibitors: crystal structures and phage display selection.,Droge MJ, Boersma YL, van Pouderoyen G, Vrenken TE, Ruggeberg CJ, Reetz MT, Dijkstra BW, Quax WJ Chembiochem. 2006 Jan;7(1):149-57. PMID:16342303 From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See Also Lipase 3D Structures

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