Conversion of the dimeric D-amino acid oxidase from Rhodotorula gracilis to a monomeric form. A rational mutagenesis approach.
The relevance of the dimeric state for the structure/function relationships of Rhodotorula gracilis D-amino acid oxidase (RgDAAO) holoenzyme has been investigated by rational mutagenesis. Deletion of 14 amino acids in a surface loop (connecting beta-strands 12 and 13) transforms RgDAAO from a dimeric protein into a stable monomer. The mutant enzyme is still catalytically competent and retains its binding with the FAD coenzyme. Dimerization has been used by this flavoenzyme in evolution to achieve maximal activity, a tighter interaction between the protein moiety and the coenzyme, and higher thermal stability.
Department of Structural and Functional Biology, University of Insubria, Via J.H. Dunant 3, 21100 Varese, Italy. luciano.piubelli@uninsubria.it
Luciano
L
Piubelli
Laura
L
Caldinelli
Gianluca
G
Molla
Mirella S
MS
Pilone
Loredano
L
Pollegioni
D000595:Amino Acid Sequence; D001483:Base Sequence; D002850:Chromatography, Gel; D003605:D-Amino-Acid Oxidase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019281:Dimerization; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009838:Oligodeoxyribonucleotides; D017433:Protein Structure, Secondary; D012248:Rhodotorula / Q000201:enzymology*; D016415:Sequence Alignment; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid
D009838:Oligodeoxyribonucleotides; D003605:D-Amino-Acid Oxidase