Biomaterial Database

Directed Evolution of Quorum-Quenching Enzymes: A Method for the Construction of a Directed Evolution Platform and Characterization of a Quorum-Quenching Lactonase from Geobacillus kaustophilus. 2018

Maybelle Kho Go, and Jeng Yeong Chow, and Wen Shan Yew

Synthetic Biology for Clinical and Technological Innovation (SynCTI), Centre for Life Sciences, National University of Singapore, Level 2, 28 Medical Drive, Singapore, 117456, Singapore.

A thermostable quorum-quenching lactonase from Geobacillus kaustophilus (GKL) was used as a template for in vitro directed evolution experiments. Here we describe the overexpression and purification of wild-type GKL, the construction of a quorum-quenching directed evolution platform using bioluminescence as a reporter, and the in vitro kinetic assay for the determination of kinetic parameters of wild-type GKL and its mutants.

UI	MeSH Term	Description	Entries
D007700	Kinetics	The rate dynamics in chemical or physical systems.
D010957	Plasmids	Extrachromosomal, usually CIRCULAR DNA molecules that are self-replicating and transferable from one organism to another. They are found in a variety of bacterial, archaeal, fungal, algal, and plant species. They are used in GENETIC ENGINEERING as CLONING VECTORS.	Episomes,Episome,Plasmid
D002265	Carboxylic Ester Hydrolases	Enzymes which catalyze the hydrolysis of carboxylic acid esters with the formation of an alcohol and a carboxylic acid anion.	Carboxylesterases,Ester Hydrolases, Carboxylic,Hydrolases, Carboxylic Ester
D003001	Cloning, Molecular	The insertion of recombinant DNA molecules from prokaryotic and/or eukaryotic sources into a replicating vehicle, such as a plasmid or virus vector, and the introduction of the resultant hybrid molecules into recipient cells without altering the viability of those cells.	Molecular Cloning
D005798	Genes, Bacterial	The functional hereditary units of BACTERIA.	Bacterial Gene,Bacterial Genes,Gene, Bacterial
D053038	Quorum Sensing	A phenomenon where microorganisms communicate and coordinate their behavior by the accumulation of signaling molecules. A reaction occurs when a substance accumulates to a sufficient concentration. This is most commonly seen in bacteria.	Quorum Quenching,Quenching, Quorum,Quenchings, Quorum,Quorum Quenchings,Sensing, Quorum
D056506	Geobacillus	A genus of gram-positive, endospore-forming, thermophilic bacteria in the family BACILLACEAE.
D057075	Enzyme Assays	Methods used to measure the relative activity of a specific enzyme or its concentration in solution. Typically an enzyme substrate is added to a buffer solution containing enzyme and the rate of conversion of substrate to product is measured under controlled conditions. Many classical enzymatic assay methods involve the use of synthetic colorimetric substrates and measuring the reaction rates using a spectrophotometer.	Enzymatic Assays,Indirect Enzymatic Assays,Indirect Enzyme Assays,Assay, Enzymatic,Assay, Enzyme,Assay, Indirect Enzymatic,Assay, Indirect Enzyme,Assays, Enzymatic,Assays, Enzyme,Assays, Indirect Enzymatic,Assays, Indirect Enzyme,Enzymatic Assay,Enzymatic Assay, Indirect,Enzymatic Assays, Indirect,Enzyme Assay,Enzyme Assay, Indirect,Enzyme Assays, Indirect,Indirect Enzymatic Assay,Indirect Enzyme Assay
D019020	Directed Molecular Evolution	The techniques used to produce molecules exhibiting properties that conform to the demands of the experimenter. These techniques combine methods of generating structural changes with methods of selection. They are also used to examine proposed mechanisms of evolution under in vitro selection conditions.	Evolution, Molecular, Directed,Molecular Evolution, Directed,In Vitro Molecular Evolution,Laboratory Molecular Evolution,Directed Molecular Evolutions,Evolution, Directed Molecular,Evolution, Laboratory Molecular,Evolutions, Directed Molecular,Evolutions, Laboratory Molecular,Laboratory Molecular Evolutions,Molecular Evolution, Laboratory,Molecular Evolutions, Directed,Molecular Evolutions, Laboratory