Metabolic Engineering of for -Amino-Phenylethanol and -Amino-Phenylacetic Acid Biosynthesis.

Aromatic amines are an important class of chemicals which are used as building blocks for the synthesis of polymers and pharmaceuticals. In this study we establish a pathway for the biosynthesis of the aromatic amines amino-phenylethanol (PAPE) and amino-phenylacetic acid (4-APA) in . We combined a synthetic -amino-l-phenylalanine pathway with the fungal Ehrlich pathway. Therefore, we overexpressed the heterologous genes encoding 4-amino-4-deoxychorismate synthase ( from ), 4-amino-4-deoxychorismate mutase and 4-amino-4-deoxyprephenate dehydrogenase ( and from ) and ThDP-dependent keto-acid decarboxylase (10 from ) in . The resulting -amino-phenylacetaldehyde either was reduced to PAPE or oxidized to 4-APA. The wild type strain LJ110 with a plasmid carrying these four genes produced (in shake flask cultures) 11 ± 1.5 mg l of PAPE from glucose (4.5 g l). By the additional cloning and expression of (phenylacetaldehyde dehydrogenase from ) 36 ± 5 mg l of 4-APA were obtained from 4.5 g l glucose. Competing reactions, such as the genes for aminotransferases ( and ) or for biosynthesis of L-phenylalanine and L-tyrosine () and for the regulator TyrR were removed. Additionally, the genes were cloned and expressed from a second plasmid. The best producer strains of showed improved formation of PAPE and 4-APA, respectively. Plasmid-borne expression of an aldehyde reductase ( from ) gave best values for PAPE production, whereas -overexpression led to best values for 4-APA. In fed-batch cultivation, the best producer strains achieved 2.5 ± 0.15 g l of PAPE from glucose (11% C mol mol-1 glucose) and 3.4 ± 0.3 g l of 4-APA (17% C mol mol glucose), respectively which are the highest values for recombinant strains reported so far.