Improved thermostability of the North American firefly luciferase: saturation mutagenesis at position 354

Autor: James A. H. Murray, Peter John White, David James Squirrell, Christopher R. Lowe, Phillipe Arnaud
Přispěvatelé: Evolution et Diversité Biologique (EDB), Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche pour le Développement (IRD)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées, Génétique, Reproduction et Développement (GReD), Centre National de la Recherche Scientifique (CNRS)-Université Clermont Auvergne [2017-2020] (UCA [2017-2020])-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut de Recherche pour le Développement (IRD)-Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Clermont Auvergne [2017-2020] (UCA [2017-2020])-Centre National de la Recherche Scientifique (CNRS)
Jazyk: angličtina
Rok vydání: 1996
Předmět:
MESH: Mutation
Mutant
Mutagenesis (molecular biology technique)
Glutamic Acid
[SDV.CAN]Life Sciences [q-bio]/Cancer
medicine.disease_cause
Biochemistry
MESH: Recombinant Proteins
[SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry
Molecular Biology/Genomics [q-bio.GN]
medicine
Photinus pyralis
MESH: Coleoptera
Animals
Luciferase
MESH: Animals
MESH: Lysine
Saturated mutagenesis
Luciferases
Molecular Biology
Thermostability
Mutation
biology
Lysine
Temperature
[SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry
Molecular Biology/Molecular biology
[SDV.BBM.MN]Life Sciences [q-bio]/Biochemistry
Molecular Biology/Molecular Networks [q-bio.MN]
Cell Biology
MESH: Glutamic Acid
biology.organism_classification
Recombinant Proteins
MESH: Temperature
Coleoptera
MESH: Mutagenesis
Site-Directed
Directed mutagenesis
Mutagenesis
Site-Directed
MESH: Luciferases
Research Article
Zdroj: Biochemical Journal
Biochemical Journal, Portland Press, 1996, pp.343-50
Biochemical Journal, 1996, pp.343-50
Scopus-Elsevier
ISSN: 0264-6021
1470-8728
Popis: International audience; We have used random chemical mutagenesis and a simple genetic screen to generate and isolate a thermostable mutant of luciferase from the North American firefly (Photinus pyralis). A single G-to-A transition mutation, resulting in the substitution of a glutamate for a lysine residue at position 354 in the protein sequence, was shown to be responsible for this enhanced thermostability. Replacement of Glu-354 with all possible amino acid residues was achieved using directed mutagenesis, and produced mutant enzymes with a range of thermostabilities. The mutations E354K and E354R conferred the largest increases in thermostability, suggesting that side-chain size and hydrophobicity, as well as charge, may also be important contributors to the overall thermostability of the polypeptide chain at this position. Unusually for such mutations, biochemical studies suggest that this position is on the surface of the protein and exposed to solvent.
Databáze: OpenAIRE
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