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Altering the Substrate Specificity and Enantioselectivity of Phenylacetone Monooxygenase by Structure‐Inspired Enzyme Redesign
Author(s) -
Pazmiño Daniel E. Torres,
Snajdrova Radka,
Rial Daniela V.,
Mihovilovic Marko D.,
Fraaije Marco W.
Publication year - 2007
Publication title -
advanced synthesis and catalysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.541
H-Index - 155
eISSN - 1615-4169
pISSN - 1615-4150
DOI - 10.1002/adsc.200700045
Subject(s) - monooxygenase , chemistry , biocatalysis , active site , substrate (aquarium) , mutagenesis , saturated mutagenesis , stereochemistry , thermostability , directed evolution , enzyme , indole test , mutant , organic chemistry , biochemistry , reaction mechanism , catalysis , cytochrome p450 , oceanography , gene , geology
Of all presently available Baeyer–Villiger monooxygenases, phenylacetone monooxygenase (PAMO) is the only representative for which a structure has been determined. While it is an attractive biocatalyst because of its thermostability, it is only active with a limited number of substrates. By means of a comparison of the PAMO structure and a modeled structure of the sequence‐related cyclopentanone monooxygenase, several active‐site residues were selected for a mutagenesis study in order to alter the substrate specificity. The M446G PAMO mutant was found to be active with a number of aromatic ketones, amines and sulfides for which wild‐type PAMO shows no activity. An interesting finding was that the mutant is able to convert indole into indigo blue: a reaction that has never been reported before for a Baeyer–Villiger monooxygenase. In addition to an altered substrate specificity, the enantioselectivity towards several sulfides was dramatically improved. This newly designed Baeyer–Villiger monooxygenase extends the scope of oxidation reactions feasible with these atypical monooxygenases.