Site Directed Mutagenesis Of Dienelactone Hydrolase

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The role of individual amino acid residues of the enzyme dienelactone hydrolase was investigated. Using the polymerase chain reaction (PCR), a 1.9 kbp clcD fragment was amplified and subcloned yielding a 821 bp BamHI to EcoRI clcD subclone in the plasmid pUC19. Site-specific mutants of dienelactone hydrolase were created using mismatched oligonucleotides to prime DNA synthesis. Specifically modified proteins from mutated clcD genes (Arg 81 to alanine, Tyr 85 to phenylalanine and Arg 206 to alanine), were encoded by the mutant clones. Enzyme assays showed that dienelactone hydrolase activity of the mutants Arg 81 and Arg 206 was totally abolished. ... continued below

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v, 72 leaves: ill.

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Chen, Wei, 1965- December 1992.

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  • Chen, Wei, 1965-

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The role of individual amino acid residues of the enzyme dienelactone hydrolase was investigated. Using the polymerase chain reaction (PCR), a 1.9 kbp clcD fragment was amplified and subcloned yielding a 821 bp BamHI to EcoRI clcD subclone in the plasmid pUC19. Site-specific mutants of dienelactone hydrolase were created using mismatched oligonucleotides to prime DNA synthesis. Specifically modified proteins from mutated clcD genes (Arg 81 to alanine, Tyr 85 to phenylalanine and Arg 206 to alanine), were encoded by the mutant clones. Enzyme assays showed that dienelactone hydrolase activity of the mutants Arg 81 and Arg 206 was totally abolished. The DLHase enzyme activity of mutant Tyr 85 is greatly decreased by approximately two thirds.

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v, 72 leaves: ill.

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  • December 1992

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  • March 9, 2015, 8:15 a.m.

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  • Aug. 28, 2017, 11:35 a.m.

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Citations, Rights, Re-Use

Chen, Wei, 1965-. Site Directed Mutagenesis Of Dienelactone Hydrolase, thesis, December 1992; Denton, Texas. (digital.library.unt.edu/ark:/67531/metadc500900/: accessed October 17, 2017), University of North Texas Libraries, Digital Library, digital.library.unt.edu; .