Coupled Natural Fusion Enzymes in a Novel Biocatalytic Cascade Convert Fatty Acids to Amines.

Tambjamine YP1 is a pyrrole-containing natural product. Analysis of the enzymes encoded in the Pseudoalteromonas tunicata " tam " biosynthetic gene cluster (BGC) identified a unique di-domain biocatalyst ( Pt TamH). Sequence and bioinformatic analysis predicts that Pt TamH comprises an N-terminal, pyridoxal 5'-phosphate (PLP)-dependent transaminase (TA) domain fused to a NADH-dependent C-terminal thioester reductase (TR) domain. Spectroscopic and chemical analysis revealed that the TA domain binds PLP, utilizes l-Glu as an amine donor, accepts a range of fatty aldehydes (C 7 -C 14 with a preference for C 12 ), and produces the corresponding amines. The previously characterized Pt TamA from the "tam" BGC is an ATP-dependent, di-domain enzyme comprising a class I adenylation domain fused to an acyl carrier protein (ACP). Since recombinant Pt TamA catalyzes the activation and thioesterification of C 12 acid to the holo -ACP domain, we hypothesized that C 12 ACP is the natural substrate for Pt TamH. Pt TamA and Pt TamH were successfully coupled together in a biocatalytic cascade that converts fatty acids (FAs) to amines in one pot. Moreover, a structural model of Pt TamH provides insights into how the TA and TR domains are organized. This work not only characterizes the formation of the tambjamine YP1 tail but also suggests that Pt TamA and Pt TamH could be useful biocatalysts for FA to amine functional group conversion.