title: Enzyme-directed mutasynthesis: a combined experimental and theoretical approach to substrate recognition of a polyketide synthase.
abstract: Acyltransferase domains control the extender unit recognition in Polyketide Synthases (PKS) and thereby the side-chain diversity of the resulting natural products. The enzyme engineering strategy presented here allows the alteration of the acyltransferase substrate profile to enable an engineered biosynthesis of natural product derivatives through the incorporation of a synthetic malonic acid thioester. Experimental sequence-function correlations combined with computational modeling revealed the origins of substrate recognition in these PKS domains and enabled a targeted mutagenesis. We show how a single point mutation was able to direct the incorporation of a malonic acid building block with a non-native functional group into erythromycin. This approach, introduced here as enzyme-directed mutasynthesis, opens a new field of possibilities beyond the state of the art for the combination of organic chemistry and biosynthesis toward natural product analogues.
authors.affiliation: Fakultät für Chemie, Chemische Biologie,  Technische Universität Dortmund , Otto-Hahn-Str. 6, 44221 Dortmund, Germany.
authors.forename: Uschi
authors.initials: U
authors.lastname: Sundermann
authors.forename: Kenny
authors.initials: K
authors.lastname: Bravo-Rodriguez
authors.forename: Stephan
authors.initials: S
authors.lastname: Klopries
authors.forename: Susanna
authors.initials: S
authors.lastname: Kushnir
authors.forename: Hansel
authors.initials: H
authors.lastname: Gomez
authors.forename: Elsa
authors.initials: E
authors.lastname: Sanchez-Garcia
authors.forename: Frank
authors.initials: F
authors.lastname: Schulz
mesh_terms: D000217:Acyltransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001688:Biological Products / Q000737:chemistry / Q000378:metabolism; D004917:Erythromycin / Q000378:metabolism; D008314:Malonates / Q000378:metabolism; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D015394:Molecular Structure; D017354:Point Mutation; D048630:Polyketide Synthases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D015202:Protein Engineering; D016930:Saccharopolyspora / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D001688:Biological Products; D008314:Malonates; D004917:Erythromycin; D048630:Polyketide Synthases; C030290:malonic acid; D000217:Acyltransferases