4u2c

Crystal structure of dienelactone hydrolase A-6 variant (S7T, A24V, Q35H, F38L, Q110L, C123S, Y145C, E199G and S208G) at 1.95 A resolution

spinqualitylabelsall models 4u2c, resolution 1.95Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image   Print3D Structural highlights 4u2c is a 1 chain structure with sequence from Pseudomonas knackmussii. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance. Method: X-ray diffraction, Resolution 1.95Å Ligands: CSD, SO4 Resources: FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT Function CLCD_PSEKB Ring cleavage of cyclic ester dienelactone to produce maleylacetate. Publication Abstract from PubMed The ease with which enzymes can be adapted from their native roles and engineered to function specifically for industrial or commercial applications is crucial to enabling enzyme technology to advance beyond its current state. Directed evolution is a powerful tool for engineering enzymes with improved physical and catalytic properties and can be used to evolve enzymes where lack of structural information may thwart the use of rational design. In this study, we take the versatile and diverse alpha/beta hydrolase fold framework, in the form of dienelactone hydrolase, and evolve it over three unique sequential evolutions with a total of 14 rounds of screening to generate a series of enzyme variants. The native enzyme has a low level of promiscuous activity toward p-nitrophenyl acetate but almost undetectable activity toward larger p-nitrophenyl esters. Using p-nitrophenyl acetate as an evolutionary intermediate, we have generated variants with altered specificity and catalytic activity up to 3 orders of magnitude higher than the native enzyme toward the larger nonphysiological p-nitrophenyl ester substrates. Several variants also possess increased stability resulting from the multidimensional approach to screening. Crystal structure analysis and substrate docking show how the enzyme active site changes over the course of the evolutions as either a direct or an indirect result of mutations. Directed Evolution of New and Improved Enzyme Functions Using an Evolutionary Intermediate and Multidirectional Search.,Porter JL, Boon PL, Murray TP, Huber T, Collyer CA, Ollis DL ACS Chem Biol. 2014 Dec 5. PMID:25419863 From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.

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