Probing the substrate specificity of aminopyrrolnitrin oxygenase (PrnD) by mutational analysis.
Molecular modeling and mutational analysis (site-directed mutagenesis and saturation mutagenesis) were used to probe the molecular determinants of the substrate specificity of aminopyrrolnitrin oxygenase (PrnD) from Pseudomonas fluorescens Pf-5. There are 17 putative substrate-contacting residues, and mutations at two of the positions, positions 312 and 277, could modulate the enzyme substrate specificity separately or in combination. Interestingly, several of the mutants obtained exhibited higher catalytic efficiency (approximately two- to sevenfold higher) with the physiological substrate aminopyrrolnitrin than the wild-type enzyme exhibited.
Lee
Jung-Kul
JK
Department of Chemical and Biomolecular Engineering, University of Illinois, 600 South Mathews Avenue, Urbana, IL 61801, USA.
Ang
Ee-Lui
EL
Zhao
Huimin
H
D000588:Amines / Q000378:metabolism*; D001665:Binding Sites / Q000235:genetics; D008958:Models, Molecular; D010105:Oxygenases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017354:Point Mutation; D011551:Pseudomonas fluorescens / Q000201:enzymology*; D011764:Pyrrolnitrin / Q000378:metabolism*; D013379:Substrate Specificity
D000588:Amines; D010105:Oxygenases; D011764:Pyrrolnitrin